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Journal of Plant Development Sciences (An International Monthly Refereed Research Journal)

Volume 8 Number 8 August 2016

Contents

RESEARCH ARTICLES

Copper and cadmium sulphide nanoparticles can induce macromutation in Nigella sativa L. (black cumin)

—Divya Vishambhar Kumbhakar, Animesh Kumar Datta, Debadrito Das, Bapi Ghosh, Ankita

Pramanik1 and Aditi Saha ---------------------------------------------------------------------------------------------- 371-377

An amino acid sequences based computational analysis of enzyme cytidylate kinase

—Nitin Kumar Verma, Balwinder Singh, Vibha ----------------------------------------------------------------- 379-383

Phytochemical screening and antibacterial activity of Elaeocarpus genitrus (Rudraksha) seeds

—Jyoti Goldar and Rituraj Gupta ----------------------------------------------------------------------------------- 385-388

Assessment of yield losses and screening of pea cultivars for resistance to root rot of pea caused Byfusarium

solani f.sp. pisi

—Anita Sharma and R.S. Ratnoo ------------------------------------------------------------------------------------ 389-393

Effect of crop establishment method and irrigation schedules on productivity and water use of wheat

—Vipin Kumar Sagar; R.K.Naresh; R.B. Yadav; Satendra Kumar; Kamal Khilari and

Raghuvir Singh ----------------------------------------------------------------------------------------------------------- 395-400

Bio-efficacy of azoxystrobin 11% + tebuconazole 18.3% sc on onion in Andhra Pradesh

—C. Ruth and M. Tagore Naik --------------------------------------------------------------------------------------- 401-404

Assessment of honey dew excretion by non -target bph, Nilaparvata lugens stal.ondifferent ir-64 bt rice events

—Gajendra Kumar, Shanjay Sharma, G. Chandel and Randeep Kumar Kushwaha------------------- 405-408

Antifungal activity of some medicinal plant extracts against human pathogenic fungus Aspergillus niger

—Arun Kumar, Vijai Malik and Shruti Saini --------------------------------------------------------------------- 409-411

Efficacy of bio-agents and organic amendments against Sclerotium rolfsii causing collar rot of Chickpea

—Santosh Lahre, N. Khare and Tikendra Kumar --------------------------------------------------------------- 414-416

Insect pests complex associated with basmati rice with western plain zone of Uttar Pradesh, India

—Kaushlendra Kumar, S.K. Sachan and D.V. Singh ----------------------------------------------------------- 417-420

SHORT COMMUNICATION

Plants as a source of diuretic activity and study of 3-(6-arylimidazo[2,1-b]thiazol-3-yl)-2-methylchromone

system as diuretic agent

—Vinay Prabha Sharma ------------------------------------------------------------------------------------------------ 421-422

*Corresponding Author

________________________________________________ Journal of Plant Development Sciences Vol. 8 (8) : 371-377. 2016

COPPER AND CADMIUM SULPHIDE NANOPARTICLES CAN INDUCE

MACROMUTATION IN NIGELLA SATIVA L. (BLACK CUMIN)

Divya Vishambhar Kumbhakar1, Animesh Kumar Datta

1*, Debadrito Das

1, Bapi Ghosh

1,

Ankita Pramanik1 and Aditi Saha

2

1Department of Botany, Cytogenetics, Genetics and Plant Breeding Section, University of Kalyani,

Kalyani - 741235, West Bengal, India 2Department of Botany, NarasinhaDutt College, Howrah 711101

Email: [email protected]

Received-12.08.2016, Revised-26.08.2016

Abstract: Dry seeds(moisture content: 5.0%) of Nigella sativa L. (Family: Ranunculaceae; common name- black cumin,

spice of commerce with immense therapeutic uses) are exposed to chemically synthesized copper (Cu) and cadmium

sulphide (CdS) nanoparticles (NPs) at the doses of 0.25, 0.50 and 1.00 µg/ml for 3 and 6 h durations . EMS (ethyl

methanesulphonate ̶ 0.25, 0.50 and 1.00%, 3 and 6 h durations) and gamma irradiations (25, 50, 100, 200 and 300 Gy; 60Co

source) are used as positive control. The objective of the work is to foresee whether NPs can induce stable phenotypic

mutation. The present communication highlights macromutation types and frequency, mutagenic efficiency and

effectiveness and meiotic chromosome behaviour in treated materials and suggests the efficacy if NPs in inducing mutation

in N.sativa and crop improvement.

Keywords: Cu- and CdS-NPs, Macromutants, Meiotic analysis, Mutagenic efficiency and effectiveness, Nigella sativa

INTRODUCTION

he nanoparticles (NPs) are characterized by their

small size and large surface and large surface

area (ranging between 1 and 100 nm ̶ Roco 2003)

and possess unique physico-chemical properties

(Remédioset al. 2012; Masarovičová and Králová

2013). NPs has global significance due to its wide

application in industry, medicine, biotechnology,

agriculture, different aspect of life sciences among

others (Roco 2003, Lam et al. 2004, Caruthers et al.

2007, Nowack and Bucheli 2007, Scrinis and Lyons

2007, Singh et al. 2008, Nair et al. 2010, Castiglione

et al. 2011,Remédioset al. 2012). Most significantly,

Halderet al. (2015 a, b) reported that chemically

synthesized copper (Cu) and cadmium sulphide

(CdS) NPs can induce stable heritable changes

(macromutation) in Macrotylomauniflorum (Lam.)

Verdc (Family: Leguminosae). Furthermore,

Kumbhakaret al. (2016) highlighted the potentiality

of Cu- and CdS-NPs as mutagenic agents in Nigella

sativa L. (Family: Ranunculaceae; common name-

black cumin) as a test material. Such findings trigger

the essentiality to attain further scientific knowledge

from plant system on induced mutagenesis following

NPs treatment. With the view to it, the present

investigation has been designed and describes the

mutagenic efficiency, mutagenic effectiveness and

macromutation types, frequencies and their meiotic

chromosome behaviour in N. sativaL. (spice yielding

plant of commerce with immense therapeutic uses-

Dattaet al. 2012) following treatments with

chemically synthesized copper and cadmium

sulphide nanoparticles in comparison with the

conventional mutagens namely, ethyl

methanesulphonate (EMS) and gamma irradiation.

The objective of the present study is to assess

whether NPs can induce similar genetic variations as

that of the well established mutagens under study.

MATERIAL AND METHOD

Germplasm

Seed samples of Nigella sativa L. (Ranunculaceae)

were collected from Medicinal Plant Garden,

NarendrapurRamkrishna Mission, Government of

West Bengal, India. The moisture content of the

mother seed stock was determined as 5.0%.

Synthesis and characterization of NPs

Cu- and CdS-NPs were prepared using wet chemical

co-precipitation methods as described earlier by

Chatterjee et al. (2012) and Halderet al. (2015 b)

respectively.The NPs were characterized using UV ̶

visible spectra (UV-vis), Fourier transform infra-red

spectroscopy (FTIR), X-ray diffraction (XRD),

Dynamic Light Scattering (DLS), Field emission

scanning electron microscopy (FESEM) and

Transmission electron microscopy (TEM)for

assessment of their nature and size (Kumbhakaret al.

2016). Opto-physical studies of the prepared NPs

confirm nano standard quality (Kumbhakaret al.

2016).

Treatments

Dry and filled seeds of N. sativa were exposed to the

prepared solutions of Cu- and CdS-NPs (0.25, 0.50

and 1.00 µg/ml, 3 and 6 h durations) and EMS (0.25,

0.50 and 1.00 %, 3 and 6 h durations; solution

prepared in 0.2 M phosphate buffer, pH 6.8). Dry

seeds were also exposed to gamma irradiation doses

(25, 50, 100, 200 and 300 Gy, source 60

Co,

T

RESEARCH ARTICLE

mailto:[email protected]

372 DIVYA VISHAMBHAR KUMBHAKAR, ANIMESH KUMAR DATTA, DEBADRITO DAS, BAPI GHOSH,

ANKITA PRAMANIK AND ADITI SAHA

absorbance dose rate 47.4 Gy/min, source to distance

12 cm). Dry control and bulk Cu- and CdS controls

(0.25 µg/ml, 3 h) were also kept for assessment. Each

lot of treatment comprised of 100 seeds.

Assessment of biological damages

Biological damages (M1 generation attributes) like

lethality, injury and sterility were ascertained from

seed germination, seedling growth and seed yield per

plant respectively. The extent of lethality and injury

were determined from the relative reduction of seed

germination frequency and seedling growth in treated

samples (under controlled Petri plate conditions- as

was suggested by Konzaket al. 1965) as compared to

controls (per cent of control). Seed sterility was

assessed in each treatment (seeds of each plant was

weighed on harvest) and is represented as percentage

of reduction in seed weight in treatment in relation to

controls (Dubey and Datta 2014).

Raising of M1 and M2 plant population

Fifty seeds from each treatment including controls

were sown in the experimental field plots of

Department of Botany, University of Kalyani (West

Bengal plains, Nadia, latitude 22˚50΄ to 24˚11΄ N,

longitude 88˚09΄ E to 88˚48΄ E, elevation 48 ft above

sea level, sandy loamy soil, pH 6.85) in late

November to raise M1 plant population. The plants

were harvested in mid-April.

Selfed seeds (first formed flower was bagged in each

case) of each surviving M1 plants were harvested

separately and were used to raise M2 (plant to row)

generation. Control lines were also grown. Plants

were grown having 15 cm between plants and 25 cm

between rows. No fertilizer was applied during any

stage of plant growth (both at M1 and M2).

Screening of phenotypic (Macromutants) mutants

Macromutants were carefully screened at M2 from

seedling to maturity and the frequency of the mutants

was estimated as per 100 plants in accordance with

Gaul (1964). Chlorophyll mutants were classified

after Blixt 1961. Seedling colors were laid down with

reference to RAL COLOUR CHART (UK). The

mutant trait(s) were confirmed at M3 from selfed

segregation of M2 mutants.

Mutagenic efficiency and effectiveness

The efficiency and effectiveness of NPs, EMS and

gamma irradiations were calculated from viable

(total) mutation frequency (Walther 1969) as

proposed by Konzaket al. (1965). The mutagenic

efficiency was calculated as Mf/L, Mf/I and Mf/S

and the effectiveness as (Mf/c) × t and Mf/Gy unit

converted to kR (Mf = mutation frequency, L =

lethality, I = injury, S = sterility, c = concentration, t

= duration, Gy = gray unit and kR = kiloroentgen).

Meiosis

Meiotic analysis was performed in controls and in

mutant plant types. For the purpose, floral buds (2 to

3 in each case) of suitable sizes were fixed (5 a.m. to

6.30 a.m.) in acetic alcohol 1:3 for overnight and

preserved in 70% alcohol under refrigeration. Pollen

mother cells (PMCs) and pollen grains obtained from

anther squash preparations were stained in 2%

acetocarmine solution. Fully stained pollen grains

were considered fertile (Marks 1954). Scattered

metaphase (MI) and anaphase I (AI) cells were only

scored for analyses. Photomicrographs were taken

from suitable preparations.

RESULT AND DISCUSSION

Mutation types andfrequencies

In comparison to normal trait(s) (Fig 1a), a total of

14 macromutant types (Table 1) are screened at M2

mutagenized population (3357 plants scored). Out of

14 types, 9 in EMS, 6 in gamma irradiations, 13 in

Cu-NPs and 14 in CdS-NPs are spotted. The mutants

comprise of two non-viable types namely,

chloroxantha (the plants dried at flowering stage: 51-

57 days from sowing, bright yellowish green colored

seedlings ̶ Fig. 1b) and viridis (medium green colored

seedlings, slow growing with reduced height, dried at

flowering stage). The viable mutants aresparsely

arranged pinnae I (associated with broad elongated

pinnae ̶ Fig. 1c) and II (Fig. 2d ̶ e ), narrow pinnae,

crumpled pinnae (upward folding of pinnae to form

cup like structure), feathery leaf, heterophyllous leaf

(broad and narrow pinnae both present ̶ Fig. 1f),

cluster pinnae top, long petiole (Fig. 1g), thick stem

(with associated bushy trait ̶ Fig. 1h), dwarf, early

flowering (37 to 39 days from sowing compared to

47 to 60 days in control plants) and synchronous

maturity (Fig. 1i). Of all the mutants, synchronous

maturity plant type is most significant as it will

minimize seed loss at harvest. This mutant type

appeared only in NPs treatments. The mutant cluster

pinnae top is found specific to CdS-NPs treatments.

The predominant mutant plant types recorded in all

treated materials are heterophyllous leaf, long

petiole, sparsely arranged pinnae I and II and

narrow pinnae. No mutant was scored in controls.

The mutant trait(s) in comparison to normal is

presented in Table 2.

Across doses of treatments it seems that EMS has

induced higher mutation frequency (viable: 9.95%,

total: 11.44% than gamma irradiations (viable:

6.32%, total: 6.32%), Cu-NPs (viable: 5.95%, total:

6.44%) and CdS-NPs (viable: 3.92%, total: 4.26%).

Higher mutation frequency in EMS than other

treating agents may be due to low number of M2

plant scored. Total failure of germination is recorded

in 1.00% EMS, 3 and 6 h durations. Over the M2

population, the macromutants (Table 1) occurred in

the following order: EMS ̶ sparsely arranged pinnae

II >chloroxantha = long petiole = dwarf>early

JOURNAL OF PLANT DEVELOPMENT SCIENCES VOL. 8 (8) 373

flowering = narrow pinnae = sparsely arranged

pinnae I >heterophyllous leaf = thick stem; gamma

irradiation ̶ heterophyllous leaf>sparsely arranged

pinnae II >long petiole>dwarf = narrow pinnae =

sparsely arranged pinnae I; Cu-NPs ̶ sparsely

arranged pinnae II>heterophyllous leaf =

synchronous maturity>dwarf>narrow pinnae = long

petiole>sparsely arranged pinnae I = feathery

leaf>viridis>chloroxantha = crumpled pinnae = thick

stem = early flowering; CdS-NPs ̶ sparsely arranged

pinnae II >narrow pinnae >heterophyllous leaf

>crumpled pinnae = cluster pinnae top = early

flowering>sparsely arranged pinnae I =

dwarf>viridis = feathery leaf = long petiole>thick

stem>synchronous maturity>chloroxantha. Apart

from EMS (0.50%, 6 h ̶ 16.67%), maximum mutation

(viable) frequency is mostly found in initial doses of

gamma irradiations (25Gy ̶ 8.82% and 50 Gy ̶

6.61%), Cu-NPs (0.50 µg/ml, 3 h ̶11.11%; 0.50

µg/ml, 6 h ̶ 16.37% and CdS-NPs (0.25 µg/ml, 3 h ̶

4.33%). The M2 mutants segregated at M3 in

accordance with Mendelian patterns (data not given

in the present communication) suggesting that the

altered trait(s) than normal are genetically controlled.

Mutagenic effectiveness and efficiency

The mutagenic effectiveness (Tables 3 and 4) relates

doses to mutation and is found to be maximum in

EMS ̶ 0.50%, 6 h, 25 Gy gamma irradiations, 0.50

µg/ml, 3 and 6 h Cu-NPs, 0.25µg/ml, 3 h CdS-NPs

treatments.

On comparative basis it can be suggested that NPs

are effective as mutagens like EMS and gamma

irradiations. The mutagenic efficiency defined as the

relation of number of mutational events to

undesirable effects (lethality, injury and sterility) and

is found to vary in relation to treating agents, and it

seems that mostly threshold doses are efficient.

Meiotic analysis and pollen grains fertility

Meiotic chromosome behaviour is nearly normal and

comparable in controls as well as in

macromutants,PMCs regularly show 2n=12

chromosomes (Fig. 2 a ̶ f) always. Controls show 6II

formation in 100% meiocytesand it varies from

97.78% (chloroxantha) to 58.28% (thick stem) in

mutants. Most of the univalents scored are rather due

to precocious separation of chromosomes as mostly

AI cells in mutants are balanced (92.45% to 100.0%).

Rarely 1 to 2 laggards are observed at AI (Fig. 2g ̶ h)

and AII (Fig. 2i). Pollen grain fertility is assessed in

controls (dry ̶ 95.80%; bulk Cu ̶ 95.79% and bulk

CdS ̶ 89.87%) as well as in mutants (70.25% in

viridis to 89.81% in heterophyllous leaf).

The present investigation reveals the potentiality of

Cu- and CdS-NPs in inducing macromutation in N.

Sativa, which is corroborating to earlier reports in

Macrotylomauniflorum (Halderet al. 2015 a, b).

Thus, NPs inducing genetic changes can be explored

for crop improvement.

Table 1. Macromutant types and frequency across doses in treatments.

Mutant types Frequency (%)

EMS Gamma irradiations CdS-NPs Cu-NPs

Chloroxantha 1.49 0.00 0.10 0.20

Viridis 0.00 0.00 0.24 0.29

Sparsely arranged pinnae I 0.99 0.55 0.34 0.39

Sparsely arranged pinnae II 1.99 1.65 0.63 1.37

Narrow pinnae 0.99 0.55 0.53 0.49

Crumpled pinnae 0.00 0.00 0.39 0.20

Feathery leaf 0.00 0.00 0.24 0.39

Heterophyllous leaf 0.50 2.20 0.48 0.78

Cluster pinnae top 0.00 0.00 0.39 0.00

Long petiole 1.49 0.82 0.24 0.49

Thick stem 0.50 0.00 0.19 0.20

Dwarf 1.49 0.55 0.34 0.68

Early flowering 0.99 0.00 0.39 0.20

Synchronous maturity 0.00 0.00 0.15 0.78

Total plants scored 201 364 2067 1025

Table 2. Mutant trait(s) in comparison to normal.

Mutants Attributes t-value DF Probability

Sparsely arranged pinnae I

(associated trait)

Interpinnae distance

Control-2.17±0.053

Mutant-3.49±0.110

Broad and elongated pinnae

Length:

control-2.22±0.029

mutant-3.06±0.040

Width:

10.77

18.78

5.52

16

18

16

>0.001

>0.001

>0.001



control-0.23±0.017

mutant-0.36±0.018

Sparsely arranged pinnae II

Interpinnae distance

Control-2.16±0.047

mutant-2.83±0.128

4.90 16 >0.001

Narrow pinnae Pinnae

Length:

control-2.22±0.290

mutant-0.76±0.037

Width:

control-0.23±0.016

mutant-0.16±0.018

29.68

2.97

18

16

>0.001

>0.01

Long petiole Petiole length

Control-5.62±0.084

Mutant-14.04±0.318

24.99 20 >0.001

Thick stem Stem thickness

Control-0.21±0.023

mutant-0.52±0.025

9.12 18 >0.001

Dwarf Height

Control-31.38±1.632

mutant-4.45±0.132

16.45 18 >0.001

Table 3. Mutagenic efficiency and effectiveness of EMS and gamma irradiation treatments in M2 generation. Treatments

(%/Gy)

Duration

(h)

Per cent of control Viable

Mutation

frequency (%)

(Mf)

Mutagenic effectiveness

(Mf/conc.)×duration

(Mf/kR)

Mutagenic efficiency

Lethality

(L)

Injury

(I)

Sterility

(S)

Mf/L Mf/I Mf/S

EM

S

0.25 3 13.89 34.38 77.41 9.09 12.12 0.26 0.15 0.36

0.50 3 13.89 50.91 75.51 8.77 5.84 0.11 0.05 0.07

1.0 3 33.33 73.82 ̶ ̶ ̶ ̶ ̶ ̶

0.25 6 8.33 41.87 75.04 6.67 2.73 0.74 0.16 0.18

0.50 6 22.22 59.93 82.46 16.67 0.54 ̶ 0.36 0.03

1.0 6 94.44 94.16 ̶ ̶ ̶ ̶ ̶ ̶

Ga

mm

a i

ra

dia

tio

ns 25 ̶ ̶ 10.07 44.08 8.82 3.53 ̶ 0.88 0.20

50 ̶ 22.22 6.91 67.14 6.61 1.76 0.30 0.96 0.09

100 ̶ 5.56 32.34 78.36 4.62 0.46 0.83 0.14 0.06

200 ̶ 52.78 62.84 100.00 ̶ ̶ ̶ ̶ ̶

300 ̶ 83.33 84.82 100.00 ̶ ̶ ̶ ̶ ̶

Table 4. Mutagenic efficiency and effectiveness of Cu- and CdS-NPs treatments in M2 generation. Treatments

(%/Gy)

Duration

(h)

Per cent of control Viable

Mutation

frequency

(%)

(Mf)

Mutagenic

effectiveness

(Mf/conc.)×

duration

Mutagenic efficiency

Lethality

(L)

Injury

(I)

Sterility

(S)

Mf/L Mf/I Mf/S

Cd

S-N

Ps

0.25 3 16.67 28.51 11.91 4.33 5.77 0.26 0.15 0.36

0.50 3 22.22 47.29 36.86 2.55 1.70 0.11 0.05 0.07

1.0 3 8.33 0.49 17.01 5.42 1.81 0.65 11.06 0.32

0.25 6 5.56 25.04 21.93 4.09 2.73 0.74 0.16 0.18

0.50 6 ̶ 4.55 40.68 1.62 0.54 ̶ 0.36 0.03

1.0 6 13.89 42.48 41.13 9.25 1.54 0.67 0.22 0.22

Cu

-NP

s

0.25 3 13.88 11.63 16.26 1.93 2.57 0.14 0.17 0.12

0.50 3 2.77 32.09 25.35 11.11 7.41 4.01 0.35 0.44

1.0 3 13.88 14.01 32.52 1.92 0.64 0.14 0.14 0.06

0.25 6 5.55 6.31 31.47 5.70 3.80 1.03 1.03 0.18

0.50 6 2.77 ̶ 44.23 16.37 5.46 5.91 5.91 0.37

1.0 6 5.55 10.07 43.53 2.88 0.48 0.52 0.52 0.06


ACKNOWLEDGMENT

Financial support from University of Kalyani is gratefully acknowledged. The authors are thankful to the UGC-

DAE Consortium, Kolkata for gamma irradiation of seeds.

Figure plate 1 (a ̶ i) showing control (Fig. a) and macromutant plant types (b ̶ i) of N. sativa. (Fig. a) control;

(Fig. b)chloroxantha; (Fig. c)sparsely arranged pinnae I-associated trait broad and elongated pinnae; (Figs. d ̶

e)sparsely arranged pinnae II; (Fig. f)heterophyllous leaf; (Fig. g)long petiole; (Fig. h)Thick stem with bushy

trait; (Fig. i) mutant showing synchronous maturity.



Figure plate 2 (a ̶ i) showing meiosis configuration in mutants at MI (a̶ ̶ f), AI (g ̶ h) and AII (i). (Figs. a ̶ d) 6II

(2n=12); (Fig. e) 1II+10I; (Fig. f) 2II+8I; (Figs. g ̶ h) 5-1-6 separation of chromosomes; (Fig. i) a laggard. Scale

bar=10µm.

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AN AMINO ACID SEQUENCES BASED COMPUTATIONAL ANALYSIS OF

ENZYME CYTIDYLATE KINASE

Nitin Kumar Verma1, 2

*, Balwinder Singh3, Vibha

4

1 Department of Biotechnology and Bioinformatics, Uttaranchal College of Science and Technology,

Dehardun, Uttarakhand, INDIA

2 Faculty of Life Science, Uttarakhand Technical University, Dehradun, Uttarakhand, INDIA

3 Department of Science, Ek Onkar Scholar Degree College, Shahbjnagar, Shahjahanpur, Uttar

Pardesh, INDIA 4 Genetics and Tree Propagation Division, Forest Research Institute, Dehradun, Uttarkhand, INDIA



Abstract: Computational analysis has been established for hypothetical study of amino acid sequences of the enzyme

cytidylate kinase that derived from various programs and databases. Cytidylate kinase enzyme is widely distributed enzyme

among bacteria and fungi. In the present study, thirteen full length amino acid sequences cytidylate kinase were retrieved,

collected and subject to multiple sequence alignment (MSA), regular expression identification, domain identification,

discovering individual amino acid composition, and construction of phylogenetic trees. Multiple sequence alignment

revealed that three glycine, one lycine, one arginine and one valine were identically found in all the bacterial and fungal

sources of cytidylate kinase. The two major sequence clusters were constructed by phylogenetic analysis. One cluster

contains two species of fungi and six species of bacteria, where as other contain five species of only fungi. The amino acid

composition results revealed that the average frequency of amino acid leucine is 9.29 % in fungi, where as alanine 13.61 %

in bacteria. In addition, six unique motifs were also identified in the group analysis.

Keywords: Motif, Phylogentic analysis, Multiple sequence alignment, Cytidylate Kinase, Domain

INTRODUCTION

inases are a universal group of enzymes, which

participate in a variety of cellular pathways. The

name kinase is applied for enzymes, which catalyze

the transfer of the terminal phosphate group from

ATP to an acceptor that can be a small molecule,

lipid, and protein substrate. The cellular and

physiological roles of kinases are different (Cheek et

al., 2002). Many kinases participate in signal

transduction pathways, in which these enzymes are

necessary components (Blenis, 1993). Other kinases

are involved centrally in the metabolism of

carbohydrates, lipids, nucleotides, amino acid

residues, vitamins, and cofactors. Some kinases play

roles in various other processes, such as gene

regulation, muscle contraction, and antibiotic

resistance. Their universal roles in cellular processes,

kinases are the best-studied enzymes at the structural,

biochemical, and cellular level. Although all kinases

catalyze essentially the same phosphoryl transfer

reaction, and display significant diversity in their

structures, substrate specificity, and number of

pathways in which they participate (Cheek et al.,

2002).

Nucleoside monophosphate kinases (NMP kinases)

are the key enzymes, which are involved in the

metabolism of nucleotides. They act specifically on

the various NMPs formed in de novo or salvage

pathways of purine or pyrimidine nucleotides, by

catalyzing the reversible transfer of a phosphoryl

group from a nucleoside triphosphate to an NMP

(Briozzo et al., 1998). CMP kinase is the key enzyme

in the nucleotide metabolism that is connected to the

family of nucleoside monophosphate kinase (NMK)

(Leipe et al., 2003). Substrate specificity was studied

on recombinant human UMP/CMP kinase

(pyrimidine nucleoside polyphosphate kinase), which

show that UMP and CMP (Verma et al., 2013) are far

better substrates than dCMP (Liou et al., 2002).

Computational methods are used to analyze protein

function that can be divided into three vast

categories: sequence, expression and interaction

based methods (Pellegrini, 2001). The success of

computational approaches is used for solving

important problems such as sequence alignment and

comparisons (Altschul et al., 1990). The importance

of this approach in research is used to annotate the

proteome through functional and structural genomic

efforts (Michalovich, 2002). Considering the above

facts, a study of amino acid sequences of cytidylate

kinase from different sources of organisms is really

challenging. In the present study, the individual

computational studies of amino acid sequences were

performed, which were obtained from bacteria, fungi,

and correlated them on the basis of some common

feature.

MATERIAL AND METHOD

The full-length amino acid sequences of cytidylate

kinase from bacteria and fungi were retrieved from

protein databases available at NCBI (National Center

for Biotechnology Information). The sequences were

K

RESEARCH ARTICLE


380 NITIN KUMAR VERMA, BALWINDER SINGH, VIBHA

arranged as in bacterial and fungal profile,

respectively. The multiple sequence alignment of the

individual profiles was performed using

CLUSTRALX (Bateman, 2007). Motifs were

discovered in profiles using the expectation

maximization approach implemented in multiple EM

for motif elicitation server (Bailey et al., 2006).

Further, the discovered motifs were used to search

their protein family using Pfam at the sanger institute

(Finn et al., 2010). The neighbor joining approach

implemented in the MEGA (Molecular Evolutionary

Genetics Analysis) program was employed for

phylogenetic analysis (Tamura et al., 2011). The

statistical reliability of the phylogenetic tree was

tested by bootstrap analyses with 500 replications.

MEGA program is also used for discovering

individual amino acid composition.


Sequence retrieval and analysis All the sequences belong to different families of

bacteria and fungi were retrieved from genbank

(National Center for Biotechnology Information)

protein database and listed in Table 1 along with their

accession number, organism name, family and

source.

Table 1. list of retrieved sequences with their different sources

S.No. Source Name of Organisms Family Accession no.

1 Bacteria Streptococcus pneumoniae Streptococcaceae KGI36288.1

2 Bacteria Staphylococcus aureus Staphylococcaceae KII20889.1

3 Bacteria Salmonella enteric Enterobacteriaceae CBY95005.1

4 Bacteria Escherichia coli Enterobacteriaceae ACA78315.1

5 Bacteria Pseudomonas aeruginosa Pseudomonadaceae KFL11511.1

6 Bacteria Mycobacterium tuberculosis Mycobacteriaceae NP_216228.1

7 Fungi Trichoderma gamsii Hypocreaceae KUE96443.1

8 Fungi Moesziomyces antarcticus Ustilaginaceae XP_014653432.1

9 Fungi Fusarium langsethiae Nectriaceae KPA47046.1

10 Fungi Rhizoctonia solani Ceratobasidiaceae CUA68089.1

11 Fungi Puccinia sorghi Pucciniaceae KNZ52000.1

12 Fungi Rhizopus microspores Mucoraceae CEG68485.1

13 Fungi Mucor ambiguous Mucoraceae GAN07890.1

Multiple sequence alignment Multiple Sequence Alignment (MSA) showed the

presence of some conserved residues in all the

sequences from different sources while others were

restricted only to their groups. Three glycine, one

lycine, one arginine and one valine were found to be

identically conserved residues in all analysed species

in bacterial and fungal profile (Figure 1 and 2).

Conserved motif identification Six conserved motifs were identified after the

analysis of bacterial and fungal profile individually.

Three conserved motifs were observed in bacterial

and fungal profile (Table 2).

Conserveed motif family identification The six identified conserved motifs were applied to

their family identification in Pfam database using

sequence search option. First three conserve motifs

identified in bacterial profile belong to

Cytidylate_kin domain family while last three

conserved motifs, a single conserved motif identified

in fungal profile belong to AAA_17 domain family

and in the Pfam entry of rest two conserved motifs

no significant family was found (Table 2).

Table 2. Motif identified using MEME program and their pfam analysis using pfam database

S.No. Motif Width Present in number

of sequences Family Sources

1

[PG]G[IL][VI][AM]DGRD[IM]GTVV[FL]P

DAP[LV]KIFL[DT]AS[ASV]EERA[EHR]R

R[YM][LK]Q[LN]Q[AE]KG[FI][ES]V[DN]

49 6 Cytidylate_kin Bacteria


FE

2

[PI]VI[AT]IDGP[AS]GAGK[GS]T[VL][AC]

K[AR][LM]A[ER][AE]L[GQ]WH[LY]LD[S

T]GA[MI]YR[AV]L[AT][LY]AAL[HK]H[G

H]VD


3

L[LK]A[ED]I[KR][EA]RDDRD[SR]NR[AE

]V[AS]PL[KV]PA[AD]DA[VL]VLD[ST]TG

[LM]SIE[EQ]V[VI]EK[IA]L[AQ]Y[AV][ER

][KQR][KR]


4

[SMP][KS][KD][ILV][FT][VR][IV][FA][VI]

[LD]G[GP]P[GA][AS]GK[GS]T[QT][CA][A

K][RL]L[VA]E[DE][YL]GF[TV][HY][LI][S

D][AS]G[DA][LM][LF]RA[EI][QT]Q[RK][

ECP][GQ][SQ]QY

50

7

AAA_17

Fungi

5

[CT][PST]E[ED][VK][ML][LE][SKP]RL[LI

]ERGKTSGR[ET]DDN[EAI]ESI[KR]KRF[

RQ]TF[VAI][EQ]TSMPV

41 7 Pfam hit not found Fungi

6

[FI]L[IVL]DGFPR[KER][ML][DE]QA[IVQ]

[KA]F[DE][EAR][ETS][VFI][CQV][PEIM][

SAP][AKQSV][FL]VLF[FL]

29 7 Pfam hit not found Fungi

Clustral analysis

Clustral analysis of bacterial profile Clustral analysis of bacteria showed two major

clusters as shown in Figure 4. Clustral A consist of

four species namely Salmonella enterica,

Escherichia coli, Pseudomonas aeruginosa and

Mycobacterium tuberculosis. Clustral B consist of

two species namely Streptococcus pneumoniae and

Staphylococcus aureus, respectively.

Clustral analysis of fungal profile Clustral analysis of fungi showed two major clusters

as shown in Figure 3. Clustral A consist of five

species namely Moesziomyces antarcticus, Puccinia

sorghi, Rhizoctonia solani, Fusarium langsethiae and

Trichoderma gamsii. Clustral B consist of two

species namely Rhizopus microsporus and Mucor

ambiguous, respectively.

Clustral analysis of joint bacterial and fungal

profiles Two major clusters were obtained by clustral analysis

of joint bacterial and fungal profiles (Figure 5).

Clustral A consists of eight species, which were

further divided into two subclustral. Subclustral A

contains three species of bacteria and two species of

fungi. Subclustral B contains three species of

bacteria and five species of fungi.

Fig. 1. Amino acid composition in different domain (Bacteria and Fungi)


Fig. 2. The conservation study of enzyme cytidylate kinase between bacteria and fungi

Fig. 3. Phylogenetic tree of fungal profile using neighbor joining method

Fig. 4. Phylogenetic tree of bacterial profile using neighbor joining method

Fig. 5. Phylogenetic tree of joint profile of bacteria and fungi using neighbor joining method


CONCLUSION

Computational analysis of the cytidylate kinase

sequences showed sequence-based similarities

depending on their source organism. Three glycine,

one valine, one lysine and one arginine residues were

identically conserved in all analyzed species. These

results suggested that the conserved amino acid

residues have an important function in cytidylate

kinase sequences. Six domains were identified in this

research work, but three domains belong to

Cytidylate_kin family that were found in all analyzed

sequences of bacteria and fungi. These domains were

found to be responsible for the functional activity of

cytidylate kinase enzyme in different source

organisms. These domains were conserved during the

evolution of lower organism and their existence is

important for the functional activity of this enzyme.

In all species of bacteria and fungi an average

frequency of amino acid leucine is 9.29 % in fungi,

where as alanine 13.61 % in bacteria, which was

higher in comparison to other amino acid average

frequency. The amino acid alanine and leucine play

an important role in the composition of cytidylate

kinase. Two major sequences cluster were obtained

by phylogenetic analysis. These phylogenetic

analysis results suggested classification significance,

which contributes the understanding of evolutionary

relationship between the species at molecular level.

Competing interests The authors declare no funding for this project, and

no competing interests exits.

ACKNOWLEDGEMENT

Authors are grateful to Director, UCST, Dehradun,

Uttarakhand, India for his generous laboratory

support and throughout inspiration for

accomplishment of this study.

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Altschul, S.F., Gish, W., Miller, W., Myers, E.W.,

Lipman, D.J. (1990). Basic local alignment search

tool. J. Mol. Biol. 215, 403–410.

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*Corresponding Author ________________________________________________

Journal of Plant Development Sciences Vol. 8 (8) : 385-388. 2016

PHYTOCHEMICAL SCREENING AND ANTIBACTERIAL ACTIVITY OF

ELAEOCARPUS GENITRUS (RUDRAKSHA) SEEDS

Jyoti Goldar and Rituraj Gupta*

Department of Biotechnology, University Teaching Department,Sarguja University Ambikapur (C.G.)



Abstract: In the present investigation phytochemical and antibacterial activity of Elaeocarpusganitrus (Rudraksha) seeds

were studied with the methanolic and acetonic extract. The major phytochemical constituents screened were tannins,

flavonoids, steroids, reducing compounds carbohydrates and alkaloids, alcohol and protein. It has been observed that

maximum phytochemical compounds were present in methanolic and acetonic extract of E. ganitrus.The phytochemical

screening was done to as certain the presence of bioactive components present in selected plant extract.Antibacterial activity

in terms of minimum inhibitory concentration (MIC) of the extracts was studied with paper disc diffusion method and zone

of inhibition was measured in mm. It has been observed that MIC was ranging from 11.25-21.25 mm for methanolic and

15.5-22mm for acetonic extract respectively. It is concluded that Rudkraksha seeds have many useful phytochemicals and

possess significant antifungal/antibacterial activity.

Keywords: Phytochemical screening, Antibacterial activity, Rudraksha

INTRODUCTION

laeocarpusganitrusis commonly known as

Rudraksha (Asolkar and Kakkar, 1992.). The

word Rudraksha literally derived from two Sanskrit

words ‘rudra’ a synonym for Lord Shiva and ‘aksha’

meaning eyes (Ramadurai, 2008). It is also called

blueberry beads as beads are covered by an outer

shell of blue color on fully ripening (Pandey and

Das, 2004)). The seed is borne by several species of

Elaeocarpus, with Elaeocarpus ganitrus being the

principal species. Rudraksha was found in tropical

and subtropical regions at the eminence ranging from

seacoast to 2,000 meters above the sea level.

Rudraksha cultivate in the area of the Gangetic plain

in the foothills of the Himalayas to South-East Asia,

Indonesia, New Guinea to Australia, Guam and

Hawaii. Rudraksha tree flourish on mountains and

hilly region of Nepal, Indonesia, Java, Sumatra and

Burma.Elaeocarpus consists of about 12 genera and

350 species of tree (Gruissem and Jones, 2000).With

the development of modern science, many scientists

researched for evidences that support the ancient

belief on the significance of Rudraksha(NISC,

2001). All the scientists came up with the findings

that reassured and confirmed the divine power of

Rudraksha beads. Individual from every walk of life

irrespective of caste, creed, religion, nationality or

gender can use Rudraksha to gain maximum

spiritual, physical and materialistic benefits.

The presence of alkaloids, carbohydrates, phenol,

tannin, flavonoids, and protein was tested by a

standard qualitative analysis (Middleton and

Kandaswami, 1994). In Indian scenario the

phytochemical screening and antibacterial activity of

Rudrakshaseeds are not known or little work has

been done. Therefore, the present study was carried

out to fill the knowledge gap in this regard.

MATERIAL AND METHOD

Rudhraksha seeds were collected from the Nitza

Biological Research Lab Hyderabad, Andhra

Pradesh, India. Epicarp and endocarp powder of

seeds were used for phytochemical analysis and

antibacterial activity as per the method reported by

Sharma and Sharma (2010).

Fig. 1. Rudraksha seeds and powder

E

RESEARCH ARTICLE


386 JYOTI GOLDAR AND RITURAJ GUPTA

Phytochemical screening

Test for alkaloids (Mayer’s Test)

Rudraksha endocarp (methanol and acetone extract)

were treated with few drop of potassium mercurric

iodide solution. Formation of a yellow colour

precipitate indicates the presence of alkaloid. And

Rudrasha epicarp (methanol and acetone extract)

were treated with few drop of potassium mercurric

iodide solution. Formation of a yellow colour

precipitate indicates the presence of alkaloid.

Test for flavonoid


were treated with sodium hydroxide solution.

Formation of intense yellow colour indicate the

absent of flavanoid.and Rudrasha epicarp (methanol

and acetone extract) were treated with sodium

hydroxide solution. Formation of intense yellow

colour indicate the absent of flavonoid (Obasiet al.,

2010; Auduet al., 2007).

Test for carbohydrates (Molisch’s Test)


were treated with few drop of alpha napthanol

solution.not formation of violet ring at the junction

indicate the absence of carbohydrates. And Rudrasha

epicarp (methanol and acetone extract) were treated

with few drop of alpha napthanol solution. Not

formation of violet ring at the junction indicates the

absence of carbohydrates.

Test for tannin


were treated with 2% gelatin solution containing

sodium chloride add the a few drops formation of

white precipitation indicate the presence of tannin

and rudrasha epicarp (methanol and acetone extract)

were treated with 2% gelatin solution containing

sodium chloride add the a few drops formation of

white precipitation indicate the presence of tannin

(Obasiet al., 2010; Audu et al., 2007).

Test for steroid


were treated with acetic anhydride and 10 ml filtrate

chloroform solution add the a few drops formation a

ring of blue-green colour indicate the presence of

steroid and Rudrasha epicarp (methanol and acetone

extract) were treated with acetic anhydroide and 10

ml filtrate chloroform solution add the a few drops

formation a ring of blue-green colour indicate the

presence of steroid (Obasiet al., 2010,Auduet al.,

2007).

Test for protein


were treated with a few drops of concentrate nitric

acid formation of yellow colour indicate the presence

of protein .and Rudrasha epicarp (methanol and

acetone extract) were treated with a few drops of

concentrate nitric acid formation of yellow colour

indicate the presence of protein (Obasi et al., 2010;

Audu et al., 2007).

Test for phenol


were treated with a few drops of ferric chloride

solution formation of bluish black colour indicates

the present of phenol. Rudraksha epicarp (methanol

and acetone extract) were treated with a few drops

of ferric chloride solution formation of bluish black

colour indicates the present of phenol (Obasi et al.,

2010; Audu et al., 2007).

Antibacterial activity

The agar well diffusion method was adopted to

assess the antibacterial activity of the selected

Rudraksha seeds (Baur et al., 1966). Further 80 ml of

nutrient agar media (NAM) was poured in four Petri

plates. The Petri plates were labeled respectively and

the media was allowed to solidified then 80µl of each

of the bacterial suspension was inoculated on the

surface of the solidified NAM.Then 80µl of extract

was added to each well respectively. The plates were

incubated at 37oC for 18 to 24 hours. After the

incubation the diameters of the growth inhibition

zone were measured as describedby Singh and Nath

(1999).


Phytochemical screening

The phytochemical screening of the rudrasha seeds

(endocarp and epicarp) revealed that alkaloid is

present in large amounts (Table no.1). And other

classes present in small quantities: tannin, steroid,

protein, phenol. However, the seed extracts tested

negative for the presence of flavanoid and

carbohydrates.

Table 1. Phytochemical constituents of Rudrasha seeds extracts

S.

No.

Family of compound Seed endocarp Seed epicarp

Methanol Acetone Methanol Acetone

1 Alkaloid +ve +ve +ve +ve

2 Flavanoid -ve -ve -ve -ve

3 Carbohydrates -ve -ve -ve -ve

4 Tannins +ve +ve +ve +ve


5 Steroid +ve +ve +ve +ve

6 Protein +ve +ve +ve +ve

7 Phenol +ve +ve +ve +ve

Fig 2. Phytochemical screening test

Antibacterial Activity

Out of Rudraksha seeds (endocarp and epicarp)

tested for antibacterial activity, Rudraksha seeds

(endocarp and epicarp) showed antibacterialactivity

by inhibiting more microorganisms. The results of

the antibacterial activity of Rudraksha seeds extracts

tested agent microorganism by disk diffusion method

are show in (table no-2) among the seeds extracts

screened the methanol and acetone extracts. The

Rudraksha seeds (endocarp and epicarp) showed in

antibacterial activity but not antifungal activity.

Table 2. Antibacterial activity of Rudraksha seeds extract

Organisms

Rudraksha Seed

Endocarp/ Epicarp

Extract Size of zone of inhibition

(Bacteria)

Micrococcus

Endocarp Methanol 13.5mm

Acetone 17.25mm

Epicarp Methanol 20.25mm

Acetone 21.75mm

E. Coli

Endocarp Methanol 12.75mm

Acetone 15.5mm


Acetone 21.5mm

Staphylococcus Endocarp Methanol 11.25mm

Acetone 20.5mm


Acetone 22mm

Fig. 2. Antibacterial activity endocarp Fig. 3. Measurment of zone (acetone epicarp)

(Staphylococcus)

388 JYOTI GOLDAR AND RITURAJ GUPTA

CONCLUSION

The selected Rudraksha seeds (endocarp and epicarp)

showed the source of the secondary metabolite i.e.

alkaloid, carbohydrates, phenol, tannin, flavanoid,

protein and amino acid. Rudrasha seeds play a vital

role in preventing various diseases, the Antioxidant

activities, anticancer activities, Antidiabetic

activities, Antifungal activities, Antibacterial

activities of the seeds are due to the presence of the

above mentioned secondary metabolite. Rudrasha

seeds are used for discovering and phytochemical

screening and antibacterial activity which are very

helpful for the manufacturing of new drugs. The

phytochemical screening and antibacterial activity of

the rudrasha seeds are also important and have

commercial. Interest in both research institutes and

pharmaceuticals companies for the manufacturing of

the new drugs for treatment of various diseases.

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The flavonoids, advances in research pp. 259–335.



ASSESSMENT OF YIELD LOSSES AND SCREENING OF PEA CULTIVARS FOR

RESISTANCE TO ROOT ROT OF PEA CAUSED BYFUSARIUM SOLANI F.SP. PISI

Anita Sharma* and R.S. Ratnoo

Department of Plant Pathology, Rajasthan College of Agriculture, MaharanaPratap University of

Agriculture & Technology, Udaipur 313001. Rajasthan, India



Abstract: Pea is an important legume crop widely cultivated throughout the world. Peas are grown in over 87 countries all

over the world (Mcphee, 2003), providing food for humans and feed for domestic animals (Hargrove, 1986; Hulse, 1994;

Patriarca et al., 2002).Although, peas have enormous nutritional qualities and have been considered to be the predominant

export crop in world trade, representing about 40% of the total trade in pulses (Oram and Agcaoili, 1994).

Keywords: Pea, Legume crop, Root rot, Disease

INTRODUCTION

n India, Uttar Pradesh ranks first in the pea

production with 1,532.39 thousand tonnes

(Chandreet al., 2014). In India, pea is widely grown

in area of 370 thousand million hectares, with

production of 3517 thousand million tonnes and

productivity 9.5 million tonnes/hectares. In Rajasthan

state, pea crop has acreage of 3.6 thousand ha with

annual production of 4.1 metric tonnes. The

productivity in the state is 1.13 metric tonnes/ha

(Anonymous, 2011). In Rajasthan pea is mainly

cultivated as a vegetable and covering the maximum

area and yielding maximum production in Jaipur

district while productivity is higher in Bharatpur and

Udaipur districts of Rajasthan (Anonymous,

1999).Among the soil borne diseases root rot of pea

is a major soil borne disease in pea growing areas

worldwide and is often considered to be the limiting

factor in pea production (Shehataet al., 1983). Root

rots are known to occur whenever peas are grown in

the world (Fenwick, 1969; Hagedorn, 1976;

Perssonet al., 1997). Root rot of pea caused by

Fusariumsolanif.sp. pisiis a disease of economic

importance causing considerable damage to the crop

and remains to be challenging task in terms of

management(Maheshwariand Jhooty,1983; Tu, 1986;

Kumar and Dubey, 2000).Root rot of pea caused by

Fusariumsolaniremained a major threat to successful

cultivation of pea and resulted in reduced crop yield

and ultimately caused heavy economic losses. It

caused severe damage at all stages of crop growth,

and up to 97% yield losses had been reported by Sen

and Majumdar (1974).

Among the soil borne diseases root rot of pea is a

major soil borne disease in pea growing areas

worldwide and is often considered to be the limiting

factor in pea production (Shehata et al., 1983).Root

rot may start when the plant is in the pre or post

emergence seedling stage. Death soon follows as

early infections, resulting in a poor crop stand. Root

decay generally begins on the finer feeder roots and

progresses gradually to the main tap root of the plant.

In some cases all roots are destroyed, leaving only

remnants below the attachment of the seed. Root rot

of pea is characterized by the cortical decay and a

brilliant red discoloration of vascular tissues in the

root (Lin et al., 1984).

The underground part of pea plant is damaged by the

fungus. On underground stem reddish brown sunken

lesions are formed. The root system may be

completely decayed and the plant has poor standing.

Vascular reddish discoloration is commonly

observed in diseased plants. Symptoms consist of

poor growth, yellowing and finally wilting of leaves

(Singh, 1999). Among the fungal diseases, the root

rot caused by Fusariumsolanif.sp.pisiremains to be

challenging task in terms of management. Therefore,

integrated management strategy is the better solution

to maintain plant health. These strategies include

minimum use of chemical for checking the pathogen

population, encouragement of beneficial biological

agent to reduce pathogen inoculums, modification of

cultural practices and use of resistant varieties

(Bendre and Barhate, 1998).

MATERIAL AND METHOD

Assessment of Yield Losses

The losses caused by a disease vary with host

pathogen interaction and the disease severity. Field

trials were conducted in two consecutive during rabi

(2010) and (2011) to assess germination percentage,

root rot incidence, green pod yield and reduction in

pod yield under different disease severity (generated

by different inoculums densities). This region has a

semi-arid climate. The soil of the experimental fields

is sandy-loam in texture, slightly alkaline (pH 7.9),

having low organic carbon (0.42) and electrical

conductivity (0.85 dSm-1). The experiment was

conducted using a local cultivar at three inoculums

densities viz., 50g/plot, 100g/plot and 150g/plot. Un

inoculated plots were maintained as control. The

I

RESEARCH ARTICLE


390 ANITA SHARMA AND R.S. RATNOO

inoculated plots were compared with un-inoculated

plots. The seeds were sown in 3x2 m plots, keeping

ten rows(30 cm) and 20 plants in each row, at 10 cm

plant distance. Recommended agronomical practices

for fertilizers (N-80, P-40 & K-40 Kg ha-1) and

weed management, pre- germination spray of

Atrazine at 0.5% and mechanical removal were

followed, but no fungicide was used in this trial. The

inoculum was multiplied on corn meal sand (2:1)

medium. The inoculations were done in the late

evening, followed by heavy irrigation to provide

adequate moisture for infection. Observation for seed

germination wererecorded at 15 days after

sowing.Theobservation for plant mortality were

recorded 90 days after sowing.

Screening of pea cultivars Eleven cultivars were evaluated under artificial

inoculation conditions using soil inoculation

technique of spore cum mycelia (50 gm/ row) of F.

solanif.sp. pisicausing root rot of pea.

Cultivars/germplasmviz., KashiSamrath, VRP-22,

VRP-6, VRP-7, VRP-5, Pea IP-3, Azad P-1, Pea KS-

210, Pea KS- 205, Pea VP-433 and Local cultivar

were evaluated against root rot pathogen in

artificially inoculated sick plots. These cultivars were

procured from IIVR, Varanasi. Seeds were sown in a

single row each of 5 m length and maintaining row

and plant to plant distance as 30x10 cm respectively.

In this screening technique a root rot susceptible

check was sown intermittently after every two test

entries so as to monitor the disease pressure.The

observations for germination percentage,% disease

incidence c.f.u. of F. solani in rhizosphere& Green

pod yield were recorded.


Assessment of Yield Losses

To determine the losses caused by root rot in pea,

field trials were conducted in the Experimental fields

of RCA, MPUAT, for two consecutive years rabi

(2010-11) and (2011-12), by creating different

disease levels through varied inoculum densities of

F. solanif.sp. pision pea local variety and compared

with uninoculated protected plots. Observations on

percent seed germination, percent plant mortality,

green pod yield and per cent yield losses were

recorded. The seed germination was recorded 15

days after sowing and plant mortality was recorded

90 days after sowing.The data are presented in Table

1.

Table 1. Effect of inoculum load of Fusariumsolanif.sp.pisi on per cent seed germination, per cent plant

mortality and per cent yield loss during rabi season (2010-11) and (2011-12) S.

No.

Treatments

(Inoculum

loads per plot

in g.)

Seed germination*

(%)

Plant mortality*

(%)

Green Pod yield*

Kg/ plot

Yield losses*

(%)

2010-11

2011-12

Pooled

2010-11

2011-12

Pooled

2010-11

2011-12

Pooled

2010-11

2011-12

Pooled

1. 50

85.77

(67.88)

87.40

(31.28)

86.59

(49.70)

20.69

(27.04)

19.53

(31.28)

20.11

(29.28)

3.35

3.38 3.37 24.73

(30.14)

21.12

(27.90)

22.92

(28.95)

2. 100 74.44

(59.26)

72.03

(32.82)

73.23

(46.12)

31.86

(34.80)

32.17

(32.82)

32.02

(33.41)

2.11 2.13 2.12 52.40

(46.60)

50.40

(45.34)

51.40

(45.95)

3. 150

68.51

(55.87)

70.55

(31.11)

69.53

(43.51)

47.70

(43.68)

45.97

(31.11)

46.84

(37.42)

1.11 1.23 1.17 74.88

(59.94)

71.09

(57.59)

72.98

(58.78)

4. Un inoculated

control

95.36

(78.16)

94.62

(30.73)

94.99

(55.60)

6.38

(14.63)

8.62

(30.73)

7.50

(24.11)

4.45 4.30 4.38 0.00

(0.00 )

0.00

(0.00 )

0.00

(0.00 )

SEm±

CD at 5%

CV%

1.22

3.75

3.73

2.15

6.61

13.33

1.23

3.60

7.16

0.49

1.50

3.25

2.15

6.61

13.33

1.10

3.21

10.02

0.11

0.34

7.90

0.05

0.17

3.89

0.06

0.18

6.22

1.42

4.37

8.30

0.73

2.26

4.48

0.80

2.33

6.75

* Mean of three replications

Figures in parenthese are arcsine per cent angular transformed values.

c.f.u of inoculum prepared in lab is 3 x 106

Pooled data of both the years (2010-11 and 2011-12)

revealed that among the different inoculum loads

used, the highest germination 86.59% recorded in 50

g inoculum/plot (T1), followed by 73.23%

germination with 100 g inoculum/plot (T2), 69.53%

germination with 150 g inoculum/plot (T3). The

highest germination 94.99% was observed in

uninoculated control plots (T4). Mean mortality

recorded in uninoculated plots (T4) was 7.50%.

Among the inoculated plots highest 46.84% mortality

was recorded with 150 g inoculum/plot (T3) followed

by 32.02% mortality with 100 g inoculum/plot (T2)

and 20.11% mortality with 50 g inoculum/plot

(T1).The uninoculated control plots (T4) yielded 4.38

kg/plot green pod yield, those inoculated with 50 g

inoculums/plot (T1) yielded 3.37 kg/plot green pod

yield, those with 100 g inoculum/plot (T2) yielded

2.12 kg/plot green pod yield and those with 150 g

inoculum/plot (T3) yielded 1.17 kg/plot green pod

yield.The highest yield loss (72.98%) among the


inoculated plots was recorded with 150 g

inoculum/plot (T3) followed by 51.40% yield loss

with 100 g inoculum/plot (T2), 22.92% yield loss

with 50 g inoculum/plot (T1).

Field experiments were conducted for two

consecutive years to estimate the losses caused by

root rot of pea with disease generated through

different inoculum densities on local variety of pea.

Results showed that the disease severity increased

with increasing of inoculum loads. A significant

reduction in green pod yield at all the severity levels

was observed as compared to the control. Pooled data

revealed that lowest germination per cent ( 69.53%)

with maximum plant mortality ( 46.84% ) and

highest yield loss ( 72.98%) was observed in plots

inoculated with higher inoculum load i.e. 150

gm/plot whereas maximum germination (86.59% )

with minimum plant mortality (20.11%) and lowest

yield loss (22.92%) was observed in plots inoculated

with low inoculums load i.e. 50 gm/plot.These

observations suggest that root rot of pea has good

potential of damaging the crops and may become

limiting factor in realization of good yield.

Screening of pea cultivars Eleven pea cultivar were screened to find out the

source of resistance against F. solanif.sp. pisi

causing root rot of pea. The experiment was

conducted in field using the varieties KashiSamrath,

VRP-22, VRP-6, VRP-7, VRP-5, Pea IP-3, Azad P-

1, Pea KS-210, Pea KS- 205, Pea VP-433 and Local

cultivar in the field where F. solanif.sp. pisiused for

soil inoculation. The experiment was conducted in

rabiseason (2010-11) and (2011-12). Observations

on germination percentage, per cent disease

incidence, green pod yield and c.f.u. of F.

solanif.sp.pisiwere recorded and presented in Table 2

. The seed germination was recorded 15 days after

sowing and plant mortality was recorded 90 days

after sowing.

Table 2. Screening of pea cultivars for resistance to root rot pathogen in field during rabi season 2010-11 and

2011-12 S.No.

Cultivar/

Germplasm

Germination*

(%)

Plant mortality*

(%)

Green pod yield*

Kg/ 5m row

Cfu of F. solanif.sp. pisi

x10490 DAS

2010-

11

2011-

12

Pooled 2010-

11

2011-

12

Pooled 2010-

11

2011-

12

Pooled 2010-

11

2011-

12

Pooled

1. Kashisamrath

82.66

(65.40)

82

(64.92)

82.33

(65.16)

46.46

(42.96)

50.59

(45.34)

48.53

(44.15) 0.81 0.71 0.76 7.8 9.3 8.55

2. VRP- 22

89.33

(70.98)

90.00

(71.62)

89.67

(71.30)

35.58

(36.61)

38.72

(38.45)

37.15

(37.53) 1.24 1.20 1.22 5.0 6.5 5.75

3. VRP- 6

92.66

(74.53)

92.00

(73.65)

92.33

(74.09)

32.11

(34.50)

33.15

(35.13)

32.63

(34.82) 1.34 1.31 1.33 3.1 3.9 3.50

4. VRP- 7

94

(75.95)

94.66

(76.70)

94.33

(76.33)

26.27

(30.80)

29.56

(32.93)

27.92

(31.86) 1.71 1.63 1.68 2.3 3.5 2.90

5. VRP- 5

91.33

(73.25)

90.66

(72.23)

91.00

(72.74)

34.29

(35.83)

36.27

(37.02)

35.28

(36.43) 1.27 1.25 1.26 4.5 5.0 4.75

6. Pea IP-3

88.65

(70.52)

88.00

(69.77)

88.33

(70.15)

39.07

(38.68)

40.43

(39.48)

39.75

(39.08) 1.14 1.10 1.12 5.5 6.8 6.15

7. Azad pea- 1

81.30

(64.39)

80.66

(63.92)

80.98

(64.16)

54.04

(47.32)

57.20

(49.21)

55.62

(48.27) 0.64 0.58 0.61 8.1 10.5 9.30

8. Pea KS-210

87.33

(69.34)

86.66

(68.73)

87.00

(69.04)

41.14

(39.89)

41.45

(40.08)

41.29

(40.12) 1.04 1.05 1.05 5.9 7.5 6.72

9. Pea KS-205

85.33

(67.55)

86.00

(68.06)

85.67

(67.81)

41.40

(40.05)

43.33

(41.16)

42.36

(40.35) 0.96 0.90 0.93 6.3 7.9 7.10

10. Pea VP-433

84.66

(67.02)

84.00

(66.45)

84.33

(66.73)

41.62

(40.17)

47.38

(43.51)

44.50

(41.78) 0.90 0.79 0.85 6.8 8.2 7.50

11. Local

79.99

(63.44)

78.00

(62.04)

79.00

(62.74)

60.30

(50.99)

59.14

(50.29)

59.72

(50.64) 0.47 0.48 0.48 8.9 10.9 9.90

SEm+ 1.45 0.89 0.85 1.03 1.44 0.89 0.04 0.045 0.03 0.27 0.29 0.20

CD 5% 4.27 2.63 2.43 3.05 4.25 2.53 0.13 0.13 0.09 0.80 0.87 0.57

CV % 4.17 2.59 3.48 5.19 7.01 6.20 8.34 9.03 8.68 9.24 8.08 8.60

* Mean of three replications

**Initial Population of F. solanif.sp. pisi 1.70 x 104c.f.u./g soil

Figures in parenthesearearcsine per cent angular transformed values. c.f.u of inoculum prepared in lab is 3 x

106


Pooled data of two seasons revealed among the

varieties, the lowest germination (79.00%) was

recorded with local pea variety followed by variety

Azad Pea-1 with 80.98% germination. KashiSamrath

with 82.33 % germination, Pea VP-433 with 84.33%

germination , Pea KS-205 with 85.67% germination,

Pea KS-210 showed 87.00% germination, Pea IP-3

with 88.33% germination, VRP-22 with 89.67%

germination, VRP-5 with 91.00% germination, VRP-

6 with 92.33% germination and the highest

germination 94.33% was recorded with VRP-7.

Among the varieties tested, the highest mortality

59.72% was recorded with local pea variety,

followed by 55.62% mortality in Azad Pea-1,

48.53% mortality in KashiSamrath, 44.50% mortality

in Pea VP-433, 42.36% mortality in Pea KS-205,

41.29% mortality in Pea KS-210, 39.75% mortality

in Pea IP-3, 37.15% mortality in VRP-22, 35.28%

mortality in VRP-5 and 32.63% mortality in VRP-6.

The lowest mortality 27.92% was recorded with

VRP-7. Highest green pod yield 1.68 kg/ row among

different varieties reported in VRP-7 as compared to

local pea which yielded lowest yield 0.48 kg/row.

With variety VRP-6 1.33 kg/row green pod yield was

reported followed by 1.26 kg/row green pod yield

from VRP-5, 1.22 kg/row green pod yield from

VRP-22, 1.12 kg/row green pod yield from Pea IP-3,

1.05 kg/row green pod yield from Pea KS-210, 0.93

kg/row green pod yield from Pea KS-205, 0.85

kg/row green pod yield from Pea VP-433, 0.76

kg/row green pod yield from KashiSamrath, 0.61

kg/row green pod yield from Azad Pea-1.

Pooled data revealed that the lowest population

density 2.90x104

cfu/g soil was observed in

rhizosphere of variety VRP-7 as compared to

rhizosphere of local pea variety where highest

population density of pathogen 9.90x104cfu/g soil

was observed. In VRP-6 rhizosphere 3.50x104cfu/g

soil was reported followed by 4.75x104cfu/g soil in

VRP-5, 5.75x104cfu/g soil in VRP-22, 6.15x10

4cfu/g

soil in Pea IP-3, 6.72x104cfu/g soil in Pea KS-210,

7.10x104cfu/g soil in Pea KS-205, 7.50x10

4cfu/g soil

in Pea VP- 433 and 8.55x104cfu/g soil in

KashiSamrath and 9.30x104cfu/g soil in Azad Pea-1.

The management of the diseases through host plant

resistance is considered as a dependable choice in all

the crop improvement programmes. Utilization of

resistance cultivars in farming is simple, effective

and economical method for management of the

diseases. The resistant cultivars reduce the cost, time

and energy when compared to the other methods of

disease management. Screening was done taking

eleven pea cultivars with inoculation of pathogen

using soil inoculation technique of spore cum

mycelia of Fusariumsolanif.sp. pisi .The result

showed that out of eleven cultivars tested one

cultivar namely VRP-7 was found moderately

resistant, four cultivars namely VRP-6, VRP-5, VRP-

22 and Pea IP-3 were found moderately susceptible

whereas six cultivars namely Pea KS-210, Pea KS-

205, Pea VP-433, Kashisamrath, Azad pea-1 and

local pea cultivar were found susceptible. Thus,

among the eleven cultivars tested none of the cultivar

was found free to the disease infection. The pooled

c.f.u. of Fusariumsolanif.sp.pisi in the rhizosphere

area of all cultivars by using serial dilution technique

at the time of sowing and 90 DAS. It was observed

that c.f.u. of pathogen in the rhizosphere of

moderately resistant cultivar was 2.90x104 whereas in

moderately susceptible cultivars it ranged from

3.50x104 to 6.15x10

4 and in susceptible cultivars it

ranged from 6.72x104 to 9.90x10

4.

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EFFECT OF CROP ESTABLISHMENT METHOD AND IRRIGATION

SCHEDULES ON PRODUCTIVITY AND WATER USE OF WHEAT

Vipin Kumar Sagar1*; R.K.Naresh

1; R.B. Yadav

1; Satendra Kumar

2; Kamal Khilari

3 and

Raghuvir Singh1

1Department of Agronomy;

2Department of Soil Science;

3Department of Plant Pathology

SardarVallabhbhai Patel University of Agriculture & Technology, Meerut-250110, U.P., India


Abstract: A field experiment was conducted during 2014-15and 2015-16 at Meerut, Uttar Pradesh. The grain yield (46.52;

47.63 and 44.01 and 44.88 q ha-1), straw (60.57; 61.55 and 59.94; 102.75 q ha-1) biological yield (107.09; 109.40 and 102.75;

104.82 q ha-1) was and harvest index (43.39; 43.49 and 42.53; 42.77) significantly higher in B90–4 and 4 cm irrigation at

IW/CPE 0.8 during both the year. Physiological traits, yield attributes and yields were significantly influenced by land

configuration and wheat irrigation schedules. In land configuration systems, B90–4and 4 cm irrigation at IW/CPE 1.2

displayed significantly higher water use efficiency (2.53; 2.51 and 2.19; 2.18 kg m-3) compared with other treatments.

However irrigation schedules × land configurationinteraction was significant for yield attributes grain, straw and biological

yield except 1000 grain weight.

Keywords: Land configuration, Irrigation schedules IW/CPE, Water use efficiency

INTRODUCTION

heat a major cereal crop is being cultivated in

the country. The main reasons for its

productivity are poor crop establishment and

improper scheduling of irrigation. Amongst the other

agronomic practices proper crop establishment

method may considerably increase the production of

wheat up to some extent. Ideal planting geometry is

important for better and efficient utilization of plant

growth resources get the optimum productivity of

wheat. It is also well know fact that water

management is one of the major factors responsible

for achieving better harvest in crop production. Both

crop establishment method and irrigation schedule

are major causes of yield reduction in wheat, which

also affect its water use efficiency. Farmers are

always interested in getting higher yield which could

not be possible without better crop management,

good stand establishment and optimum utilization of

resources. Crop production is influenced by its

establishment and plant vigor representing the key

factors towards crop development (Amanullahet al.,

2009).

To increase the water productivity of wheat,

CYMMIT introduced a planting pattern termed as

furrow irrigated raised bed planting system in

Mexico. The adoption of the system rose from 6% of

farmers in 1981 to 75% in 1994 in high-yielding

irrigated wheat-growing areas of northwestern

Mexico (Sayre and Hobbs 2004). In this system, the

crop is planted on the top of beds and irrigation water

is applied in furrows. The width of the bed and

furrows commonly used are 40–45 and 25–30 cm,

respectively, and the bed height is 15 cm– 20 cm.

Inspired by the success of irrigated maize–wheat on

permanent raised beds in Mexico, furrow irrigated

raised bed planting system was introduced in Indo-

Gangetic Plains in the mid-1990s for wheat (Sayre

and Hobbs 2004). Even after 2 decades of its

introduction and promotion, a few farmers preferred

bed planting over the conventional flat planting

system. This was mainly due to lack of yield

advantage in furrow irrigated raised bed planting

system over flat planting system. Farmers can easily

respond to this technology if efforts are diverted to

demonstrate yield differences between flat and bed

planting systems either by modifying bed

configuration or crop rows planted on the top of the

bed or selection of suitable cultivars. Keeping in

view the above points, a study was conducted to

compare crop establishment method especially in

different bed size configurations and rows planted on

the top of the bed with flat planting in wheat.

Different crop establishment method were assessed

for wheat productivity and water saving.

MATERIAL AND METHOD

Experimental site The field experiment was established in 2014 at

SardarVallabhbhai Patel University of Agriculture

&Technology, Meerut research farm (290 04', N

latitude and770 42' ‘E longitude a height of 237m

above mean sea level) U.P., India. The region has a

semi-arid sub-tropical climate with an average

annual temperature of 16.80C. The highest mean

monthly temperature (38.90C) is recorded in May,

and the lowest mean monthly temperature (4.50C) is

recorded in January. The average annual rainfall is

about 665 to 726 mm (constituting 44% of pan

evaporation) of which about 80% is received during

the monsoon period. The predominant soil at the

experimental site is classified as TypicUstochrept.

Soil samples for 0–20 cm depth at the site were

collected and tested prior to applying treatments and

W

RESEARCH ARTICLE

396 VIPIN KUMAR SAGAR; R.K.NARESH; R.B. YADAV; SATENDRA KUMAR; KAMAL KHILARI AND

RAGHUVIR SINGH

the basic properties were non-saline (EC 0.42 dS m-1

)

but mild alkaline in reaction (pH 7.98). The soil

initially had 4.1 g kg-1

of SOC and 1.29 g kg-1

of total

N (TN), 1.23 g kg-1

of total phosphorus, 17.63 g kg-1

of total potassium, 224 mg kg-1

of available N, 4.0

mg kg-1

of available phosphorus, and 97 mg kg-1

of

available potassium.

Experimental design and management

A detailed description of crop establishment methods

are necessary to compare the influence of land

configuration practices on environmental

performance (Derpsch et al., 2014).Six crop

establishment methodsB1- 75 cm bed, 2 rows (B75–2);

B2-, 75 cm bed, 3 rows (B75–3) ; B3- 90 cm bed, 2

rows (B90–2) ; B4- 90 cm bed, 3 rows (B90–3), B5- 90

cm bed, 4 rows (B90–4); B6- Flat planting , rows 22.5

cm apartin main plots and three irrigation schedule

practices were I1-4 cm irrigation at IW/CPE 0.8; I2-

5 cm irrigation at IW/CPE 1.0; I3- 6 cm irrigation at

IW/CPE 1.2allotted to sub-plots in a split-plot design

andreplicated thrice. The gross and net plot sizes

were 7.0 m×24.5 m and 6.0 m×3.5 m, respectively

and treatments were superimposed in the same plot

every year to study the cumulative effect of

treatments.

Preparation of furrow irrigated raised beds

At the beginning of the experiment soil was tilled by

harrowing and plowings followed by one field

leveling with a wooden plank, and raised beds were

made using a tractor-drawn multi crop zero till cum

raised bed planter with inclined plate seed metering

devices. The dimension of the raised beds were 45

and 60 cm wide (top of the bed) x 18 cm height x 30

cm furrow width (at top) and the spacing from center

of the furrow to another center of the furrow was

kept at 75 and 90 cm. In furrow irrigated raised bed

planting system, the crop was planted on the top of

beds in bed configurations of 45 cm bed and 60 cm

bed.

Preparation of Conventional tillage

After the rice harvest, following the conventional

practice of two harrowing, two ploughing (using a

cultivator) and one planking (using a wooden plank)

that followed pre-sowing irrigation and wheat was

seeded in flat planting, a uniform row-to-row

distance of 22.5 cm was maintained. Using a seed

drill with a dry-fertilizer attachment.

Nutrient application

Plant nutrients were applied as per the state

recommendations for wheat (N120+ P60 + K40). Urea,

di-ammonium phosphate and muriate of potash, were

placed in band in seed rows at the time of sowing

using zero till cum raised beds planter with inclined

plate metering device. The remaining N was

broadcasted with dry urea in two equal splits of 30

kg N ha–1

, (N30) at crown root initiation (CRI) and

the flag leaf initiation (FLI) crop growth stages.

Sowing techniques

Wheat cultivar DBW-17 was shown on November

2015 and 2016 using 80 kg ha-1

for raised beds and

100 kg seed ha–1

for flat planting was done using zero

till cum raised beds planter with inclined plate

metering device.

Weed management

The crop was maintained with weed free using

following practices. Weeds were controlled by

spraying of herbicide Sulfosulfuron + Metsulfuron

(Total) 35 g a.i. ha-1

and applied uniformly in

standing crop to control the weeds at 30-45 DAS. To

check the weed growth, one inter culture operation

was done during 2014-15 and 2015-16 eight weeks

after sowing with the help of manual weeding.

Irrigation scheduling

Measured quantity of irrigation water was applied to

the plots as per the irrigation schedule. For

measuring irrigation water, volume method was

used. Irrigations were scheduled on IW: CPE ratio in

individual treatments. The source of irrigation water

was Tube well with good quality water for irrigation.

Water application and measurements Irrigation water was applied using polyvinyl chloride

pipes of 15-cm diameter and the amount of water

applied to each plot was measured using a water

meter (Dasmesh Co., India). The quantity of water

applied and the depth of irrigation was computed

using the following equations:

Quantity of water applied (L) = F × t ...(1)

Depth of water applied (cm) = L /A /1000 ...(2)

Where F is flow rate (L/s), t is time (s) taken during

each irrigation and A is area of the plot (m2). Rainfall

data was recorded using a rain gauge installed within

the meteorological station. The total amount of water

(input water) applied was computed as the sum of

water received through irrigation (I) and rainfall

(R).Water productivity (WPI+R) (kg/m3) was

computed as follows (Humphreys et al, 2008)

WPI+R =Grain yield/(Irrigation water applied (I) +

Rainfall received by the crop (R)). ...(3)

Water use studies Soil moisture content was measure at seeding, and

before and after each irrigation on the top of the

ridge and furrow in furrow irrigated raised bed

planting system and between the 2 rows in flat

planting by using neutron moisture meter. Water

saving (WS) was calculated as:

WS = (QF – QB)/QF × 100,

Where QF and QB are quantity of water applied in flat

planting and furrow irrigated raised bed planting

system, respectively. The soil moisture data would be

utilized to calculate the consumptive use.



Yield attributes

Data on various yields attributing characters

viz. spike length, number of spikelet’s spike-1

,

number of grains spike-1

, and test weight, as

influenced by land configuration and different

irrigation schedules are presented in (Table 1)

revealed that B75-2 land configuration significantly

higher spike length over other treatments. The

number of spikelet’s spike-1

, number of grains

spike-1

, and test weight higher with B90-2 as

compare to remaining treatments during the year of

study. The number of grains pike-1

was higher in 90

cm than 75 cm beds and flat planting. The irrigation

scheduling 4 cm irrigation at IW/CPE 0.8 recorded

significantly values for all the above yield attributes

as compare to other irrigation schedules. Stimulated

vegetative growth of wheat on account of adequate

and prolonged supply of water in treatment

manifested itself in increased spike length, number

of spikelet’s spike-1

, number of grains spike-1

,

and test weight( Jat and singh 2003; Maurya and

singh 2008;).

Interaction effects between irrigation schedules and

land configuration in relation to spike length,

number of spikelet’s spike-1

, number of grains

spike-1

, and test weight were significant (Table 1).

The magnitude of increase in spike length due to

improvement in moisture supply by irrigation with

furrow irrigated raised beds was observed under

IW/CPE 0.8 (I1) irrigation schedules with B90-2 land

configuration as compared to other treatments

combination during 2014-15 and 2015-16,

respectively.

Table 1. Effect of land configuration and irrigation schedules on yield attributes

Yield

The grain (46.52, 47.63 q ha-1

), straw (60.57, 61.55 q

ha-1

), biological 107, 109.40 q ha-1

) yields and

harvest index (43.39 and 43.49) significantly higher

(Table 2) were recorded with B90-4 land

configuration the as compared to all other

treatments during experimentation. The grain

yield increased 11.00 and 12.02 %, straw yield

7.0 and 7.3% with B90-4land configuration over

flat planting during first and second year,

respectively. Treatments B75-4 (B2) and flat

planting (B6) were at par with each other during both

the year of study. However, B90-2 (B3) was recorded

the lowest grain yield during both the year of study.

Higher grain yield with bed planting of wheat has

been also reported by (Bhahmaet al. 2007; Kumar

2010; Thindet al. 2010).

The results have clearly shown that the grain yield in

land configurations B75–2, B75–3, B90–2 and B90–3 was

lower than that in flat planting due to low plant

density, but the yield was higher in B90–4 (B5) than

flat planting. The irrigation schedules having good

tillering and higher rates of photosynthesis, had high

biomass production and therefore was more suited

for furrow irrigated raised bed planting system than

flat planting.

Among the irrigation schedules IW/CPE 0.8 (I1) and

IW/CPE 1.0 (I2) produced higher number of spikes

Treatment

Spike length

(cm)

No. of Spikelet’s spike-1 No. of grains spike-1 Test weight

(g)

2014-15 2015-16 2014-15 2015-16 2014-15 2015-16 2014-15 2015-16

Land configuration

75 cm bed, 2 rows 12.3 12.5 20.8 22.1 54.7 56.8 43.46 43.80

75 cm bed, 3 rows 11.3 11.5 19.3 20.3 51.0 53.0 41.40 41.67

90 cm bed, 2 rows 12.1 12.3 24.2 25.2 55.9 58.6 44.15 44.67

90 cm bed, 3 rows 11.0 11.3 23.9 25.2 55.4 57.8 44.01 44.41

90 cm bed, 4 rows 11.0 11.2 21.3 22.3 52.8 54.4 43.51 43.87

Flat planting 10.1 10.3 18.8 19.8 50.0 52.3 42.74 42.95

SEm(±) 0.06 0.07 0.32 0.20 0.35 0.45 0.14 0.15

C.D. (P=0.05) 0.18 0.22 1.00 0.63 1.11 1.41 0.45 0.46

Irrigation schedules

4 cm irrigation at IW/CPE 0.8 12.1 12.2 22.7 24.1 55.0 57.7 44.17 44.58



SEm(±) 0.05 0.05 0.12 0.11 0.20 0.20 0.10 0.12

C.D. (P=0.05) 0.14 0.14 0.36 0.32 0.57 0.58 0.29 0.34

Interaction I × B Sig Sig Sig Sig Sig Sig NS NS


RAGHUVIR SINGH

and biological yield than IW/CPE 1.2 (I3). The

significantly higher grain, straw, biological yields

and harvest index was obtained in IW/CPE 0.8 (I1)

irrigation schedules and increased the grain yield

17.27 and 17.02 % over IW/CPE1.2 (I3).

Interaction effects between irrigation schedules and

land configuration in relation to grain yield, straw

yield and biological yield were significant (Table 2).

The magnitude of increase in spike length due to

improvement in moisture supply by irrigation with

furrow irrigated raised beds was observed under

IW/CPE 0.8 (I1) irrigation schedules with B90-2 land

configuration as compared to other treatments

combination during 2014-15 and 2015-16,

respectively.

Table 2. Effect of land configuration and irrigation schedules on grain, straw, biological yield and harvest index

Consumptive use

The consumptive use of water (23.0 and 23.8 cm)

was more under flat method (Table 3) followed by

the B75-2, B75-3, B90-2, B90-3 and lowest value of

consumptive use was recorded under B90-4land

configuration during both the year of study. The

consumptive use of water directly related with

moisture depletion and it washigher under flat

method and lowest under bed B90-4land

configuration. During 2015-16 total consumptive use

of water was more than 2014-15.

Treatment Consumptive Use (cm)

Water use efficiency

(kg m-3)

2014-15 2015-16 2014-15 2015-16

Land configuration

75 cm bed, 2 rows 22.2 22.8 1.77 1.76

75 cm bed, 3 rows 21.6 22.1 1.86 1.87

90 cm bed, 2 rows 19.3 20.0 1.96 1.93

90 cm bed, 3 rows 19.2 19.7 2.24 2.24

90 cm bed, 4 rows 18.4 19.0 2.53 2.51

Flat planting 23.0 23.6 1.82 1.80

Irrigation schedules

4 cm irrigation at IW/CPE 0.8 23.2 23.8 1.90 1.88



The consumptive use of water directly related with

moisture depletion and it was higher under flat

method and lowest under B90-4 (B5) land

configuration. So, consumptive use of water was also

in the order of moisture depletion. During first year

total consumptive use of water was more than the

Treatment

Grain yield

(q ha-1)

Straw yield

(q ha-1)

Biological yield

(q ha-1)

Harvest index

(%)

2014-15 2015-16 2014-15 2015-16 2014-15 2015-16 2014-15 2015-16

75 cm bed, 2 rows 39.33 40.18 55.33 56.42 39.33 40.18 55.33 56.42

75 cm bed, 3 rows 40.28 41.39 56.47 57.20 40.28 41.39 56.47 57.20

90 cm bed, 2 rows 37.80 38.57 54.63 55.36 37.80 38.57 54.63 55.36

90 cm bed, 3 rows 43.06 44.12 57.79 58.33 43.06 44.12 57.79 58.33

90 cm bed, 4 rows 46.52 47.63 60.57 61.55 46.52 47.63 60.57 61.55

Flat planting 41.92 42.52 56.59 57.35 41.92 42.52 56.59 57.35

SEm(±) 0.57 0.54 0.74 0.80 0.57 0.54 0.74 0.80

C.D. (P=0.05) 1.79 1.70 2.34 2.50 1.79 1.70 2.34 2.50




SEm(±) 0.18 0.18 0.28 0.29 0.18 0.18 0.28 0.29

C.D. (P=0.05) 0.51 0.53 0.83 0.86 0.51 0.53 0.83 0.86

Interaction I × B Sig Sig Sig Sig Sig Sig NS NS


second year mainly due to the differences in weather

conditions, such as hot and dry wind and lesser

number of rainy day.

Table 3. Effect of land configuration and irrigation

schedules on consumptive use and water-use

efficiency

Consumptive use by the crop includes total soil

moisture depletion (cm) and soil moisture contributes

Irrigation schedule of 4 cm irrigation IW: CPE 0.8.

In contrast, the lowest consumptive use of water

(17.2 and 17.6 cm) was under the irrigation schedule

of IW: CPE1.2 due to combination of higher surface

evaporation and moretranspiration so that moisture

stresses condition occurs (Ahamad 2002; Maurya

and singh 2008).

The consumptive use of water showed an increasing

trend with increase in irrigation water during both the

years. The highest consumptive use was recorded

with irrigation schedule of IW: CPE 0.8 (I1). This

was mainly due to fact that the greater loss of applied

water through evapotranspiration because of more

availability of water resulted into better foliage and

ultimately better plant growth. As a result of this was

greater absorption of moisture by crop favored by

highest water use at wettest regime. In contrast, the

lowest consumptive use of water (17.6 and 17.2 cm)

was under the irrigation schedule of IW: CPE 1.2 (I3)

due to combination of lower surface evaporation and

reduced transpiration under less moisture

availability.

Water-use efficiency

It is evident from the data (Table 3)that highest

wateruseefficiency was recorded (2.53 and 2.51kg m-

3) under B90-4 (B5) land configuration over flat

planting method (B6) during both the year of study.

Treatment B90-4 (B5) increased 39.01and 39.44%over

flatmethodduring 2014-15 and 2015-16, respectively.

This might be due to higher grain yield obtained

under B90-4(B5) land configuration with lesser amount

of water used. Declined water-use efficiency (WUE)

under flatmethod with IW: CPE 0.8 (I1) might be due

to fact that grain yield did not

increaseproportionately to that of consumptive use

under this treatment.

An examination of data (Table 3) clearly indicates

thatwater-use efficiency decreased with increase in

levels ofirrigation during both the years. Maximum

value of WUE, 2.19 and 2.18kgm-3

, were noted

inIW: CPE 1.2 (I3) during first and second

yearsrespectively. It increased 15.2 and

16.0%overIW: CPE 0.8 (I1) during first and second

year, respectively. However minimum water-use

efficiency wasunder IW: CPE 0.8 (I1) during both the

years. Decrease in WUE with IW: CPE 0.8 (I1)

basedon the fact that the proportionate increase in

grain yield wasless than increase in the consumptive

use of water.

Water productivity

The maximumwater productivity was registered

(2.45and 2.98 kg m-3

) under B90-4 land configuration,

followed by B90-3> flat, B75-3>B75-2>B90-2, treatments

during both the years.

Higher water productivity (2.21kg m-3

) was affected

by irrigation schedule of IW: CPE 1.2 (I3) during

2014-15,but during 2015-16 higher water

productivity(2.93 kg m-3

) was observed under

IW/CPE 1.0 (I2).Increase in water productivity

(Table 4) with IW: CPE 1.2 (I3) based on the fact that

the proportionate increase in grain yield with lesser

number of irrigations during experimentation(Kumar

2010; Singh et al., 2015).

In general, water productivity affectedby irrigation

schedules the higher water productivity observed

with IW: CPE 0.8 (I1) during both the year of study.

However minimum water productivity was observed

in IW: CPE 1.2 (I3).Decrease in water productivity

with IW: CPE 1.2 (I3),

Table 4. Effect of land configuration and irrigation schedules on water productivity Total water used by the

crop includes applied irrigation and effective rainfall

Treatment Total water applied (cm) Water Productivity (kg/m-3)

2014-15 2015-16 2014-15 2015-16

Land configuration

75 cm bed, 2 rows 19 16 2.07 2.51

75 cm bed, 3 rows 19 16 2.12 2.59

90 cm bed, 2 rows 19 16 1.99 2.41

90 cm bed, 3 rows 19 16 2.27 2.76

90 cm bed, 4 rows 19 16 2.45 2.98

Flat planting 19 16 2.21 2.66

Irrigation scheduling

4 cm irrigation at IW/CPE 0.8 21 16 2.10 2.81




RAGHUVIR SINGH

REFERENCES

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(2009). Timing and Rate of Phosphorus Application

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Jat, L. N. and Singh, S. M. (2003). Varietal

suitability, productivity and profitability of wheat

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under furrow-irrigated raised bed system. Indian

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Ashok (2010). Enhancing yield and water

productivity of wheat (Triticumaestivum) through

furrow irrigated raised bed system in the Indo-

Gangetic Plains of India. Indian Journal of

Agricultural Sciences; 80(3):198-202.

Maurya, R. K., Singh, G. R. (2008). Effect of crop

establishment methods and irrigation schedules on

economics of wheat (Triticumaestivum) production,

moisture depletion pattern, consumptive use and crop

water-use efficiency.Indian Journal of Agricultural

Sciences; 78(10):830-833.

Sayre, K.D. and Hobbs, P.R. (2004). The raised-

bed system of cultivation for irrigated production

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Uphoff N and Hansen D O (Eds).pp 337–55.

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Sewhag, Meena and Pannu, R.K. (2015). Water

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(TriticumaestivumL.) under varying moistureregimes

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Irrigation Sciences28(6): 489–96.



BIO-EFFICACY OF AZOXYSTROBIN 11% + TEBUCONAZOLE 18.3% SC ON

ONION IN ANDHRA PRADESH

C. Ruth* and M. Tagore Naik

Department of Plant Pathology, Dr. YSR Horticultural University, Horticultural College & Research

Institute, Anantharajupeta, Kadapa dt., Horticultural Research Station, Mahanandi - 518502,

Kurnool District, Andhra Pradesh


Abstract: Field trials conducted against Azoxystrobin 11% + Tebuconazole 18.3% SC on Onion in Andhra Pradesh.

Experimental findings with the data pertaining to efficacy of different fungicidal formulation on the purple blotch incidence

showed that all the treatments were significantly superior over control in reducing the disease severity. Azoxystrobin 11% +

Tebuconazole 18.3% SC @ 750 & 1000 ml/ha is superior and lowest disease incidence was recorded (17.15 & 18.05

respectively) and proved to be the best. Highest yield was obtained in treatment sprayed with the Azoxystrobin 11% +

Tebuconazole 18.3% SC @ 1000 ml/ha and it was on par with Azoxystrobin 11% + Tebuconazole 18.3% SC @ 750ml/ha

with the highest cost benefit ratio of 1:2.16.

Keywords: Bio-efficacy, Azoxystrobin, Tebuconazole, Purple blotch

INTRODUCTION

nion (Allium cepa L.) is a high value spice cum

bulbous vegetable crop cultivated in almost all

parts of the country. In India, onion occupies an area

of 0.52 million hectare with the production of 6.50

million tonnes. Even though India ranks first in area

under onions in the world and second in production

but its productivity is low (12.5 t/ha) as compared to

worlds productivity (Anon., 2004), (Kappa Kondal,

2014).

Onion is cultivated throughout the year in Kurnool

district, which is one of the largest producing onion

district. The area is more in Gonegandla, Kodumuru,

C. Belagal, Veldurthi, Bethamcherla, Orvakallu and

Nandikotkuru mandals. The most important varieties

are Bellary red, Agrifound light red, N-53 and Orient

hybrid. The major constraints in onion production are

spurious seed, uneven bulb development, price

fluctuations and diseases. Among several factors,

diseases are one of the most important factors

associated with low productivity in onion. Purple leaf

blotch caused by Alternaria porri is one among the

serious fungal diseases that affect onion, causing

heavy yield loss ranging from 2.5 to 87.8 per cent

during kharif season (Srivastava et al., 1994).

MATERIAL AND METHOD

A field experiment was conducted at Horticultural

Research Station, Mahanandi, Kurnool, (A. P.) to

study the efficacy of Azoxystrobin 11% +

Tebuconazole 18.3% SC in Agrifound dark red

variety of onion with 30x15cm spacing and 5x5 M

plot size during kharif 2013 and 2014. The

experiment was laid out in Randomized Block

Design (RBD) and the crop was raised by standard

agronomic practices. The test fungicide,

Azoxystrobin 11% + Tebuconazole 18.3% SC was

evaluated at 3 doses 750, 1000 and 1250 ml/ha

against purple blotch disease.

The crop was raised as per the recommended

package of practices, except plant protection

measures. The first treatment spray was done soon

after the onset of the disease and subsequent two

sprays were taken up at an interval of 15 days.

Rating scale for assessment of Purple blotch

disease

Observations were recorded at first appearance of the

disease symptoms on leaves till the harvest at weekly

intervals. The percent disease intensity was recorded by

using 0-5 scale for onion purple leaf blotch (Sharma,

1986).

The details of 0-5 scale (Sharma, 1986) in onion purple blotch

Grade Description of the symptoms

0 No disease symptom

1 A few spots towards tip covering 10 percent leaf area.

2 Several purplish brown patches covering up to 20 percent of leaf area

3 Several patches with paler outer zone covering up to 40 percent leaf area.

4 Leaf streaks covering up to 75 percent leaf area or breaking of the leaves from center

5 Complete drying of the leaves or breaking of leaves from center.

O

RESEARCH ARTICLE

402 C. RUTH AND M. TAGORE NAIK

The percent disease index of purple leaf blotch in onion was calculated using the following formula:

Percent Disease Index = Sum of the individual diseases grade x 100

(PDI) Number of leaves observed x Maximum Disease grade

Table 1. Evaluation of Azoxystrobin 11% + Tebuconazole 18.3% SC against Purple blotch in Onion in first season

*Figures in parenthesis are angular transformed value

Table 2. Evaluation of Azoxystrobin 11% + Tebuconazole 18.3% SC against Purple blotch in Onion in second

season

*Figures in parenthesis are angular transformed value

RESULT

In first season, the data pertaining to efficacy of

different fungicidal formulation on the purple blotch

incidence showed that all the treatments were

significantly superior over control in reducing the

disease severity. Azoxystrobin 11% + Tebuconazole

18.3% SC @ 750 & 1000 ml/ha is superior and

lowest disease incidence was recorded (17.26 &

16.54 respectively) and proved to be the best (Table

1). Highest yield was obtained in treatment sprayed

with the Azoxystrobin 11% + Tebuconazole 18.3%

SC @ 1000 ml/ha and it was on par with

Azoxystrobin 11% + Tebuconazole 18.3% SC @

750ml/ha with the highest cost benefit ratio of 1:2.11

In second season, recorded the same results, that all

the treatments were significantly superior over

control in reducing the disease severity.

Azoxystrobin 11% + Tebuconazole 18.3% SC @ 750

& 1000 ml/ha is superior and lowest disease

incidence was recorded (17.15 & 18.05 respectively)

and proved to be the best (Table 2). Highest yield

was obtained in treatment sprayed with the


Sl. No Treatment Formulation

ml/ha

*Purple

blotch

Onion Yield

(t/ha)

T1 Azoxystrobin 11% + Tebuconazole 18.3% SC 500 20.14 (26.64) 18.46



T4 Azoxystrobin 23% SC 500 22.05 (27.97) 19.00

T5 Tebuconazole 25.9% EC 750 23.24 (28.79) 18.05

T6 Difenconazole 25% EC 500 25.28 (30.13) 17.12

T7 Control - 29.55 (32.90) 13.80

CD 5% 1.88 1.15


ml/ha

*Purple

blotch

Onion Yield

(t/ha)




T4 Azoxystrobin 23% SC 500 22.84 (28.52) 19.80

T5 Tebuconazole 25.9% EC 750 23.66 (29.06) 18.15

T6 Difenconazole 25% EC 500 24.58 (29.67) 17.06

T7 Control - 30.25 (33.34) 14.54

CD 5% 1.50 1.21


1000 ml/ha and it was on par with Azoxystrobin 11%

+ Tebuconazole 18.3% SC @ 750ml/ha

The cost benefit ratio calculated for different

fungicides revealed the highest C:B ratio was found

in with Azoxystrobin 11% + Tebuconazole 18.3%

SC @ 750ml/ha and 1000 ml/ha (1:2.11 and 1:1.01)

respectively were found to be superior over rest of

the treatments (Table No.3).

Table 3. Economics of using Azoxystrobin 11% + Tebuconazole 18.3% SC against Purple blotch in Onion

DISCUSSION

The results were in accordance with the following

reports. .Chethan et al (2013) revealed that mancozeb

was most effective in reducing the disease intensity

by 44.43% and increasing the yield by 43.57%.

T.harzianum & garlic extract were the next best

effective treatments to reduce the disease intensity by

32.05 and 27.19% and increased the yield by 39.63

and 34.94%. The incremental cost benefit ratio was

recorded highest in mancozeb (1:14.60) followed by

T. harzianum (1:10.05). in this study showed that

mancozeb was the only fungicide found to be most

effective than bioagents and botanicals. However all

other botanicals and bioagents were at par with

fungicides in reducing the disease intensity and

increasing the yield? Thus the effective bioagents

and botanicals could be employed individually or by

integration with fungicides for the efficient

management of purple blotch disease to ensure

bioefficacy of fungicides against Alternaria porri

(Ellis) Cif., causing purple blotch of onion (Allium

cepa L.). Chethan et al (2011) studied on different

fungicides and these findings were accordance with

this new result.

Rao et al. (2015) also proved that Purple blotch

disease caused by Alternaria porri is a major

production constraint in onion, causing severe crop

loss ranging between 30 and 100%. In this study,

evaluated fungicides including new molecules for the

management of purple blotch disease of onion. 10

nonsystemic fungicides,13 systemic fungicides and 6

combination products at different concentrations

were evaluated against A. porri under in vitro

condition. Among them a nonsystemic fungicide,

Mancozeb 70% WP @ 2500 ppm and a combination

product, Cymoxanil 8%+Mancozeb 64% @ 2500

ppm were best by completely inhibiting mycelial

growth and conidial germination of A. porri.

Efath Shahnaz (2013) studied that foliar blight is an

important disease of onion, proving a major

bottleneck in its production. Six pathogens were

found associated with the disease, viz., Alternaria

alternata, A. porri, A. tenuissima, Stemphylium

vesicarium, Colletotrichum circinans and

Cladosporium alliicepae. Integrated disease

management of the crop was attempted using

chemicals (mancozeb at 0.25% and hexaconazole at

0.06%), biocontrol agents, Trichoderma viride (Tv1)

and Trichoderma harzianum (Th1), each at 1×109

spores/ml and phytoextracts (Cannabis indica and

Curcuma longa, each at 10%). Mancozeb at 0.25 per

cent proved most effective in managing foliar blight

of onion but was at par with hexaconazole at 0.06 per

cent. Among biocontrol agents used, application of

T. harzianum (Th1) resulted in lower disease

intensity as compared to T. viride (Tv1), though both

were statistically at par with each other, but were

significantly superior over the control. The

phytoextracts, C. indica and C. longa were

ineffective in the disease management.

Sobhy II Abdel-Hafez et al., investigated onion

purple leaf blotch was controlled by plant extracts

like Azadirachta indica under invitro conditions.


ml/ha

Yield

(t/ha)

Overall

income

(Rs./ha)

Economic

Benefit

over

control

T1 Azoxystrobin 11% +Tebuconazole 18.3% SC 500 18.46 97800 1:1.47

T2 Azoxystrobin 11%+Tebuconazole 18.3% SC 750 20.24 106250 1:2.11

T3 Azoxystrobin 11% +Tebuconazole 18.3% SC 1000 21.25 100700 1:2.01

T4 Azoxystrobin 23% SC 750 19.00 95000 1:1.37

T5 Tebuconazole 25.9% EC 400 18.05 85750 1:1.15

T6 Difenconazole 25% EC 500 17.12 91100 1:1.26

T7 Control - 13.80 66000 -

404 C. RUTH AND M. TAGORE NAIK

Rahman et al. (1989) proved maximum disease

reduction was highest with Rovral, Dithane M-45 in

controlling purple leaf blotch of onion.

Savitha (2015), significant effect was found with the

seed treatment using Pseudomonas fluorescens (5

g/kg) followed by two sprays of difenconazole

(0.1%) interspersed with P. fluorescens (0.5%) spray

with a per cent disease index of 38.67 and bulb yield

of 19.41 t/ha, which was significantly superior over

other treatments and control. Whereas, the seed

treatment with P. fluorescens followed by two sprays

of iprodione + carbendazim (Quintal) (0.2%)

interspersed with P. fluorescens (0.5%) spray was on

par with standard check (two sprays of difenconazole

at 0.1%) with the PDI of 42.00 and 43.33 and bulb

yield of 16.64 and 15.68 ton/ha respectively.

CONCLUSION

It is evident from the present investigation during

two seasons Kharif-2013 and Kharif-2014,


750ml/ha showed with the highest cost benefit ratio

of 1:2.11 and lowest purple leaf blotch disease

incidence 17.26.(PDI)

REFERENCES

Anonymous (2004). The Hindu Survey of Indian

Agriculture, The Hindu Publications, Chennai, pp.

120.

Rahman, L; Ahmed, H.U., Main, I. H. (1989).

Abstracts of annual review, Institute of Postgraduate

studies in Agriculture, Salna, Gazipur (Bangladesh),

1989 p.27

Srivastava, P.K., Bharadwaj, B.S. and Gupta , P.

P. (1994). Status of field diseases and selected pests

of onion in India. News letter National Horticultural

Research Developmental Found, 14(2): 11-14

Chethana, B. S., Girija, Ganeshan and

Manjunath, B. (2011). Screening of genotypes and

effect of fungicides against purple blotch of onion.

International Journal of Science and Nature, 2 (2):

384-387.

Chethana, B.S; Girija, Ganeshan, Rao, A.S. and

Bellishree (2013). Bioefficacy of botanicals,

bioagents and fungicides in the management of

purple blotch disease of onion Journal Environment

and Ecology Vol. 31 No. 2B pp. 947952

Environment and Ecology Vol. 33 No. 1 pp. 99104

Rao, A.S., Girija, Ganeshan, Ramachandra, Y. L.,

Chethana, B.S. and Bellishree, K. (2015) Journal of

Environment and Ecology 2015 Vol. 33 No. 1

pp.99104.

Efath, Shahnaz, V. K., Razdan, Rezwi, S. E. H.,

Rather, T. R., Gupta, Sachin (2013) Integrated

Disease Management of Foliar Blight Disease of

Onion: A Case Study of Application of Confounded

Factorials Journal of Agricultural sciences .volume 5

no.1 p. 17

Kappa, Kondal (2014), Growth Rate of Area,

Production and Productivity of Onion Crop in

Andhra Pradesh, Journal of applied research,

Volume-4, ISSR 2249-555 X

Savitha, A.S; Ajithkumar, K. and Ramesh, G. (2014). Integrated disease management of purple

blotch [Alternaria porri (Ellis) Cif ] of onion. Pest

management in Horticultural Ecosystems, Vol.20,

No.1pp 97-99



ASSESSMENT OF HONEY DEW EXCRETION BY NON -TARGET BPH,

NILAPARVATA LUGENS STAL.ONDIFFERENT IR-64 BT RICE EVENTS

Gajendra Kumar*1, Shanjay Sharma

1, G. Chandel

2 and Randeep Kumar Kushwaha

1

1Department of Entomology, CoA, IGKV, Raipur, Chhattisgarh, India-492012

2Department of Plant Molecular Biology & Biotechnology, CoA, IGKV, Raipur,

Chhattisgarh India-492012


Abstract: The experiment was undertaken at greenhouse of Entomology and Department of Plant molecular biology

&biotechnology, CoA, Raipur during 2014 and 2015.Area marked due to honey dew excretion by BPH under different IR64

Bt rice events ranged from 15.52 to 24.85 mm2. The maximum marked area (24.85 mm2) was observed in IR-64-C followed

by TN-1-C (23.58 mm2) with minimum in Ptb-33-C (15.52 mm2) during 2014. Whereas during 2015, new starved female

was released and new filter paper was kept inside the funnel to receive the honey dew in all the rice events were ranged from

11.72 to 20.43 mm2. The maximum marked area (20.43 mm2) was observed in IR-64-4 followed by IR-64-1 and TN-1-C

(23.58 mm2), respectively and minimum in Ptb-33-C (11.72 mm2). On the basis of two years, pooled mean of honey dew

area marked under different rice events was ranged 13.62 to 21.43 mm2. The highest honey dew excreted on IR64 Bt events

was noticed (21.43 mm2) in IR-64-4 followed by TN-1-C (20.84 mm2) and minimum in Ptb-33-C (13.62 mm2) within 24hrs.

releasing of BPH. The descending order of honey dew excretion by starved female on Bt events was as IR-64-4> TN-1-C >

IR-64-C> IR-64-1> IR-64-2> Ptb-33-C. The area of honey dew excretion by female on Bt rice and on non-transgenic control

rice plants did not differ significantly.

Keywords: Bt protein, Non-target insect BPH, Honey dew excretion

INTRODUCTION

ice is the most remunerative crop stands first

among all food grain and is staple food for more

than half of world’s population. Insect pest are one of

the major constraints of high tech agriculture and

pesticides use is necessary .The transgenic plants

expressing insecticidal properties are becoming

environmentally safe alternatives to chemical

pesticides. Genetically modified crop containing

crystal protein from the bacterium Bacillus

thuringiensis (Bt) was grown on 26.3 million ha

worldwide in the year 2005 (James, 2005). Bt rice

has the potential to eliminate yield losses caused by

lepidopteron pests up to 2%-10% of Asia’s annual

rice yield of 523 million tons (High et al., 2004).

Genetically modified crops had provided economic

benefits to growers and also offer a promising

alternative to chemical insecticides for control of

lepidopteran pests in rice (Zhu, 2001; High et al.,

2004). Zhou et al., (2004) had detected the impact

of Bt rice on non -target Nilaparvata lugens and he

did not find any difference in feeding and oviposition

behavior. In recent years, rice stem borers had

developed resistance to some most commonly used

insecticides in China and other rice growing

countries. The transgenic rice on target lepidopteran

pests is a important tool both for pest management

and insecticide resistance management. The Bt rice

has effectively controlled the three species of stem

borers (C. suppressalis, S. incertulas, S. inferens) and

leaf folder (C. medinalis) as reported by Tuet al.

(2000).The rice field has highly diverse and

interlinked insect pest species of herbivores,

predators, and parasitoids. These are an essential

component of biological control and are one of the

fundamentals of insect management strategies in rice

(Schoenly et al 1998). This importance was

previously exemplified by the outbreaks of brown

plant hopper, Nilaparvata lugens

(Homoptera:Delphacidae) that resulted from the

excessive use of insecticides in early 1997 (Gallagher

et al1994). For biological control in managing the

balance of insect pest population in rice fields, it is

essential to assess the effect of novel insecticides

such as Bt on non-target insects and their

predators/parsitoids. Mirid bug, Cyrtorrhinus

lividipennis (Hemiptera: Miridae) survives

predaciously by feeding on N. lugens larvae and

nymphs in the rice ecosystem. Insight into the

potential effect of the deployment of transgenic

Btrice on the population dynamics of other non-target

insects and their predatory organisms can be gained

by evaluating the effects of Bt toxins on life-history

parameters of brown planthopper and mirid bug

feeding on Bt rice and control non-Bt rice.

This study was undertaken to assess the effect of Bt

toxins on life-history parameters of brown plant

hopper, a non-target insect of rice to understand the

secondary exposure of Bt toxins. N. lugens is a major

pest of rice and an important constituent of the

R

RESEARCH ARTICLE

406 GAJENDRA KUMAR, SHANJAY SHARMA, G. CHANDEL AND RANDEEP KUMAR KUSHWAHA

population structure of the rice growing area of

Asian countries.

MATERIAL AND METHOD

Experimental details

The experimentalwas undertaken at glasshouse of

Entomologyand Department of Plant molecular

biology & biotechnology, College of Agriculture,

Raipur during 2014 and 2015.

Mass culture of BPH

The macropterous females of N. lugens50 to 100 per

field were collected from Entomological rice fields

of IGKV,Raipur. These females were pooled and

allowed to oviposit on caged rice plants of Taichung

Native-1 (TN-1), a rice genotype that does not

contain any genes with resistance against these pest

species. From the resulting progenies,colonies

weremaintained on susceptible rice variety TN-1.The

culture of BPH is being maintained throughout the

experimental period in the air cooled glass house in

the Department of Entomology at 30C 5C on

potted TN1 variety of rice. BPH were reared on 40 to

45 days old potted TN1 plants inside a rearing cage

of 75 x 75 x 75 cm size, consisting of wooden frame

with small window on front side and fine wire mesh

on top and other sides. Cages were mounted on

cemented platform having water level of 7.5 cm.

Potted TN1 plants were placed inside the rearing

cages for egg laying along with at least 60 pairs of

BPH per pot. After 2-3 days the females starts egg

laying inside the leaf sheath at the basal portion of

paddy plants. After the emergence of nymphs from

plants BPH pairs were transferred to another TN1

pots with the help of aspirator for egg laying.The

colonies were grown on an artificial diet before

releasing them onto transgenic Bt rice plants.The

BPH population was taken from mass culture

maintained in the glasshouse. Standard evaluation

technique developed by IRRI was adopted to

evaluate different Bt/non Bt lines. Observations were

recorded on the honey dew excretion for all the rice

events.

Collection and quantification of honeydew of N.

lugens

The parafilm sachet method as described by Pathak

et al. (1982) was used to collect honeydew from the

female adults of N. lugens fed on transgenic Btrice

and their counterpart control rice lines. Three sachets

were attached to the stems of each plant. Plants were

grown in the transgenic greenhouse arranged in a

complete block design (CBD) at 25-30C under a

natural photoperiod of approximately L12:D12.

Three fifth instar nymphs were randomly selected

from N. lugens colonies maintained on caged TN-1

rice plants and starved for 2 hrs were then placed

singly into inverted plastic cups enclosing a plant

stem and allowed to feed for 24 hrs. After 24hrs, the

filter papers were collected and the area of honeydew

spots on the filter paper was determined by placing

the transparent sheets with 1 mm grids on top of the

filter paper. Blue and white spots, produced by

alkaline and acidic honeydew deposition,

respectively, were determined separately. The

honeydew excreted was collected with the help of a

micropipette and placed into 1.5-ml micro centrifuge

tubes. The honey from each plant was pooled and

stored at -20C/-80

C. There were three replicates for

each transgenic line and three for their control

plants.The presence of Cry protein in the honeydew

secretions was analyzed using the CryIAb/CryIAc

ELISA kit (Envirologix, USA) at 450 nm as per the

manufacturer’s instructions Reading for each

replication was recorded separately. Honeydew

production on rice plants at the maximum tillering

stage was quantified using the bromocresol green

technique (Pathak and Heinrich 1982).

Table 1. Honeydew excretion by BPH on differenttransgenic rice (including control) lines during 2014 and

2015

Transgenic rice Honeydew excretion by BPH within 24h (mm

2)

2014 2015 Pooled mean

IR-64-1 18.98

(4.41)

18.10

(4.31) 18.54

IR-64 -2 19.10

(4.43)

17.89

(4.29) 18.49

IR-64-3 20.18

(4.55)

14.93

(4.93) 17.56

IR-64-4 22.42

(4.79)

20.43

(4.58) 21.43


IR-64-C 24.85

(5.03)

14.74

(4.90) 19.80

TN1-C 23.58

(4.91)

18.10

(4.31) 20.84

Ptb33-C 15.52

(4.00)

11.72

(3.50) 13.62

SEM 4.32 2.90

CD at 5% 7.57 5.09

CV (%) 36.23 30.37

*The values in parenthesis are square root transformed values

* Three replications for each treatment

Fig.1. Honeydew excretion by BPH on differenttransgenic rice (including control) lines during 2014 and 2015


Area marked due to honey dew excretionby BPH

under different IR64 Bt and non-Bt events ranged

from 15.52 to 24.85 mm2. The maximum marked

area (24.85 mm2) was observed in IR-64-C followed

by TN-1-C (23.58 mm2) with minimum in Ptb-33-C

(15.52 mm2) during 2014. Whereas during 2015, new

starved female was released and new filter paper was

kept inside the funnel to receive the honey dew in all

the treatment and replications were ranged from

11.72 to 20.43 mm2. The maximum marked area

(20.43 mm2) was observed in IR-64-4 followed by

IR-64-1 and TN-1-C (23.58 mm2), respectively and

IR-64-4 (22.42 mm2)and minimum in Ptb-33-C

(11.72mm2). On the basis of two years, pooled mean

of honey dew area marked under different treatments

was ranged 13.62 to 21.43 mm2. Maximum honey

dew excreted onIR64 Bt and non-Bt events was

noticed (21.43 mm2) in IR-64-4 followed by TN-1-C

(20.84 mm2) and minimum in Ptb-33-C (13.62 mm

2)

within 24hrs releasing of BPH (table-1 & Fig.-1).

The honeydew excreted by females of N. lugens was

quantified by measuring the area of white and blue

spots on bromocresol green-treated filter paper.

White spots, indicating the deposition of acidic

honeydew from xylem feeding, were significantly

fewer in the transgenic Bt rice lines than in the

corresponding control plants. whereas, the area of

blue spots indicating the feeding from the phloem

region was significantly higher in transgenics than in

the control rice plants. Our results clearly indicate

that BPH feeding on Bt rice has shown a preferred

feeding behavior from phloem tissues. The results

observed with the honeydew excreted by N. lugens

females feeding on Bt rice observed are similar to

those in an earlier study by Bernal et al (2002) where

toxic protein was detected in honeydew and phloem

tissues of Bt rice plants.

On the basis of honey dew area marked under

different treatments at 48hrsreleasing of BPH, it may

be stated that starved female feeding on IR-64-4Bt

plant, honey dew excretion increased gradually and it

was higher to other Bt and non- Bt control whereas,

in case ofPtb-33-C treated plant, the insect start

0

5

10

15

20

25

2014 2015

Pooled mean

Are

a o

f h

on

ey

de

w e

xcre

tio

n (

Av

mm

2)

Events

IR-64-1 IR-64 -2 IR-64-3 IR-64-4 IR-64-C TN1-C Ptb33-C

408 GAJENDRA KUMAR, SHANJAY SHARMA, G. CHANDEL AND RANDEEP KUMAR KUSHWAHA

feeding subsequent but thequantum of feeding was

low as compared to other Bt events within 24hrs.The

descending order of honey dew excretion bystarved

female on Bt events wasas IR-64-4> TN-1-C> IR-64-

C> IR-64-1> IR-64-2>Ptb-33-C. Are of honey dew

excretion by female on Bt rice and on non-transgenic

control rice plants did not differ significantly.

REFERENCES

Bernal, C. C., Aguda, R. M. and Cohen, M. B.

(2002). Effect of rice lines transformed with Bacillus

thuringiensis toxin genes on the brown plant hopper

and its predator Cyrtorhinus lividepennis.

Entomol.Exper. Appl. 102: 21-28.

Gallagher, K. D., Kenmore, P. E. and Sogawa, K.

(1994). Judicial use of insecticides deter plant hopper

outbreaks and extend the life of resistant varieties in

Southeast Asian rice. pp. 599-614. In R. F. Denno

and J. T. Perfect (eds.), Plant hopper: their ecology

and management. Chapman and Hall, New York.

High, S. M.; Cohen, M. B.; Shu, Q.Y. and

Altosaar, I. (2004). Achieving successful

deployment of Btrice. Trends in PlantScience, 9,

286-292.

James, C. (2005). Executive summary: Global Status

of Commercialized Biotech/GM Crops: 2005. ISAAA

Briefs, No. 34, ISAAA: Ithaca, NY.

Pathak, P. K. and Heinrich, E. A. (1982).

Bromocresol green indicator for measuring feeding

activity of Nilaparvata lugens on rice varieties.

Philipp. Entomol. 11: 85-90.

Pathak, P. K., Saxena, R. C. and Heinrichs, E. A.

(1982). Parafilm sachet for measuring honeydew

excretion by Nilaparvata lugens on rice. J. Econ.

Entomol.. 75: 194–195.

Schoenly, K. G., Justo, Jr. H D, Barrion, A. T.,

Harris, M. K. and Bottrell, D. G. (1998). Analysis

of invertebrate biodiversity in Philippines farmer’s

irrigated rice field. Environ. Entomol. 27: 1125-

1136.

Tu, J., Zhang, G., Datta, K., Xu, C., He, Y.,

Zhang, Q., Khush, G. S. and Datta, S. K. (2000).

Field performance of transgenic elite commercial

hybrid rice expressing Bacillus thuringiensis

endotoxin. Nat. Biotechnol. 18: 1101-1104.

Zhou, X.; Cheng, J. A.; Hu, Y. and Lou, Y.G.

(2004). Effects of transgenic Btrice on the population

development of Nephotettixcincticeps. Chinese

Journal of Rice Science, 19, 74-78.

Zhu, F.Sh.; Chen, H. X. and Lu, Y. Sh. (2001).

Outbreak reasons and management technology of

Spodoptera lituraon economic crops.Plant Prot.

Technol. Extens. 21 (7), 22.



ANTIFUNGAL ACTIVITY OF SOME MEDICINAL PLANT EXTRACTS AGAINST

HUMAN PATHOGENIC FUNGUS ASPERGILLUS NIGER

Arun Kumar*, Vijai Malik and Shruti Saini

Department of Botany M.S. College Saharanpur (U.P.) India



Abstract: The present investigation was carried out to observe the antifungal activity of Alstonia scholaris Argemone

maxicana, Datura alba, Solanum nigrum and Solanum xanthocarpum. For this purpose effect of different alcoholic extract

concentration was observed on growth performances of Aspergillus niger on 5th and 7th day. Our result shows that alcoholic

extract concentrations inhibit radial growth of this fungus. Results also indicate that inhibition of fungal growth increase

with the increase in the concentration of alcoholic extracts.

Keywords: Antifungal activity, Alcoholic extract, Aspergillus niger, Medicinal plants

INTRODUCTION

erbal medicines used against various fungal

diseases. Antifungal activity of natural plant

extract and pure compound can be detected by

inhibition of various microflora like yeast, fungi by

samples that are placed with them. About 100,000

species of fungus are present in the environment and

more than 100 of them are pathogenic in human

(Keeler 1991). Many of the Pharmaceuticals like

opium, aspirin, digitalis, quinine etc have a long

history of usage as herbal remedies. It is estimated

that about 25% of all modern medicines are directly

or indirectly derived from higher plants (Cragg et al.,

1997 and Shu, 1998). Plants and their extracts have

been used all over the world in folk medicines and

the use of extracts has been supported by the

isolation of antifungals from the plants. Many plants

produce secondary metabolites. These metabolites

may serve as potent antimicrobial agents and thus

may be useful for human beings. It has been

estimated by the World Health Organization (WHO)

that 80 percent of the population of some Asian and

African countries presently use herbal medicine for

some aspect of primary health care (Akerele, 1993).

Lupeol and Epicatechin have been identifies in the

methanol extract of Alstonia scholaris. This extract

has shown antioxidant and anticancer effect. It also

showed significant antimicrobial effect against

Staphylococcus aureus and gram negative organisms

like Escherichia coli, Proteus vulgaris, Pseudomonas

auriginosa and Candida albicans (Thara and Zuhra,

2013). Traditionally herbal medicines provide an

interesting, largely unexplored source of potential

new drugs (Udgirkar et al., 2012). Antifungal

activity of eight medicinal plants extract (Aloe vera,

Ocimum sanctum, Cenetella asiatica, Piper betle,

Calotropis gigantea, Vitex negundo, Ocimum

basilicum and Azadirachta indica) was assayed by

agar well diffusion method on plant pathogenic

fungus (red rot disease causing agent) Colletotrichum

falcatum. The result revealed that the extract of eight

medicinal plants showed significant reduction in

growth of C. falcatum (Prince and Prabakaran,

2011). Antony et al., 2012 have reported that

butanolic extract of bark of the Alstonia scholaris

have potent anti-tubercle effect and anti-

Mycobacterium tuberculosis potential and it was

concluded that it is a promise for future therapeutic

interventions. Therefore present investigation has

been aimed to evaluate the antifungal activity of

alcoholic extracts of five medicinal plants against the

pathogenic fungus viz. Aspergillus niger.

MATERIAL AND METHOD

Sample Collection

Samples for the following medicinal plants were

collected from district Saharanpur & Shiwalik belt of

Uttar Pradesh as well as from Garhwal hills of

Uttarakhand, India.

1. Alstonia scholaris

2. Argemone maxicana

3. Datura alba

4. Solanum nigrum

5. Solanum xanthocarpum

The freeze-dried pathogenic fungi Aspergillus niger

was obtained from Forest Pathology Division, Forest

Research Institute, Dehradun. The cultures were

maintained on Sabouraud Dextrose Agar (SDA)

slants and kept refrigerated until used. The SDA

plate cultures were inoculated from the slants and

incubated at 25 ± 1°C for 7 days.

Plant Extract Preparation

For the preparation of various plant extracts 5 gm of

fresh plant part was washed 2-3 times with distilled

water and then treated with 0.1% HgCl2 solution for

sterilization. After surface sterilization plant samples

were ground in mortar and pestle with 50%

methanol. The homogenized liquid was filtered and

centrifuged at 5000 rpm. The supernatants were used

as test extract & make up into 20 ml using 50%

methanol. Further, the extract was diluted into

H

RESEARCH ARTICLE


410 ARUN KUMAR, VIJAI MALIK AND SHRUTI SAINI

different concentrations, i.e. 10%, 25% and 50%. 20

ml of SDA (Sabouraud Dextroes Agar) culture

medium with 5 ml of the above concentration of the

extracts were poured in sterile petriplates and

allowed to solidify. In the control same volume of

distilled water (in place of experimental material)

was mixed in appropriate amounts.

Fungal Inoculation

For antifungal activity mycelia discs of 5 mm

diameter were cut from the periphery of 7 day old

culture of the test organisms and were aseptically

inoculated upside down on the surface of the SDA

medium in plates. Inoculated petriplates were

incubated at 250

C ± 10C and observation were

recorded at 5th

and 7th

day.

After 5th

and 7th

day of incubation, observations were

recorded on the basis of colony diameter (cm) on

medium and percent inhibition of radial growth was

calculated using following formula:

% Growth Inhibition =

Colony diameter in control

– Colony diameter in treated sets x 100

Colony diameter in control

OBSERVATION AND RESULT

The present investigation was carried out to observe

the antifungal activity of Alstonia scholaris,

Argemone maxicana, Datura alba, Solanum nigrum

and Solanum xanthocarpum. For this purpose effect

of different alcoholic extracts concentrations with

(10%, 25% and 50%) were observed on the growth

performances of Aspergillus niger causing human

skin diseases are given in Table 1.

Antifungal activity of Alstonia scholaris on

Aspergillus niger

Table 1 shows that in 10%, 25% and 50% alcoholic

root extracts of Alstonia scholaris the growth of

Aspergillus niger was 75.0%, 62.5%, 50% control

respectively in 5th

day old culture plate. Similarly,

the growth is inhibited in presence of alcoholic shoot

and seed extracts in culture medium. However, the

inhibition of growth found more at higher

concentrations. So in presence of 50% alcoholic

shoot and seed extract the growth of Aspergillus

niger is 43.3% and 48.3% of control respectively at

7th

day. Nearly similar pattern of growth inhibition

found in various other concentrations at both days of

studies.

Antifungal activity of Argemone maxicana on

Aspergillus niger

Observation shows that alcoholic extracts of

Argemone maxicana plant part is inhibitory to the

growth of Aspergillus niger. Results have shown

that higher concentration of alcoholic extract is

inhibitorier as compared to the lower concentration

of alcoholic extracts. Thus, 50% root extract causes

67% growth inhibition at 7th

day. Likewise, growth

of Aspergillus niger on both day in various

concentrations of seed extracts also inhibited. Thus,

in 50% alcoholic seed extract at 5th

day and 7th

day

the growth is 50.0% and 55.5% of control

respectively.

Table 1 also shows that shoot extract concentrations

are also inhibitory to the growth of Aspergillus niger.

Thus, in 10%, 25%, and 50% alcoholic root

concentration the fungal growth at 7th

day is 90.0%,

70.0% and 46.6% of control respectively.

Antifungal activity of Datura alba on Aspergillus

niger

Result in table 1 shows that alcoholic plant part

extracts of Datura alba are inhibitory to the growth

of Aspergillus niger. Results have shown that

inhibition of fungal growth increases with the

increase in the concentrations of alcoholic extracts.

Thus, 10% alcoholic seed extract causes 18.0%

inhibition of Aspergillus niger growth at 7th

day,

however, this inhibition in 50% alcoholic seed

extract at 7th

day is ca. 52%. Root and shoot extracts

of this plant also inhibits the growth of Aspergillus

niger in culture medium. Thus, in 10%, 25%, and

50% alcoholic root extract the Fungal mycelial

growth is 85.7%, 68.8% and 53.5% of control

respectively at 7th

day. Likewise, in 10%, 25% and

50% shoot extract concentration the mycelial growth

is 83.3%, 66.6% and 46.6% of the control

respectively at 7th

day of growth.

Antifungal activity of Solanum nigrum on

Aspergillus niger

Studies have shown that with the increase in

concentration of alcoholic extract the inhibition of

radial growth of Fungi also increases. Thus, in 10%,

25% and 50% alcoholic concentration of root extract

the growth is 85.2%, 70.50% and 50.0% respectively

of the control at 7th

day of radial growth.

Table 1 shows that like root extract growth of

Aspergillus niger is also inhibited in the various

concentrations of shoot and seed extracts. Result

shows that 50% alcoholic seed extract is inhibitory

by 42% at 7th

day of radial growth of Aspergillus

niger.

Antifungal activity of Solanum xanthocarpum on

Aspergillus niger

Result shows that Solanum xanthocarpum carries

fungicidal property to control the growth of this

fungi. Observation further shows that alcoholic

extract of various plant parts of this plant retards the

radial growth of this fungi. Thus, radial growth of

this fungi on 7th

day in presence of 10%, 25% and

50% alcoholic root extract are 80.0%, 72.0% and

52.0% of the control respectively.

Result further shows that like root extract growth of

this fungi also inhibited in the presence of shoot


extract and seed extract. Thus, in 50% alcoholic

extract concentration of shoot and seed at 7th

day the

growth is 58.3% and 57.6% of the control

respectively. Thus, the above studies shows that the

growth of this fungi affected by the alcoholic extract

of above medicinal plant.

Table 1. Antifungal activity of Alstonia scholaris, Argemone maxicana, Datura alba, Solanum nigrum,

Solanum xanthocarpum on growth performance of Aspergillus niger Days Alstonia scholaris Argemone maxicana Datura alba Solanum nigrum Solanum

xanthocarpum

Root Shoot Seed Root Shoot Seed Root Shoot Seed Root Shoot Seed Root Shoot Seed

Growth in Control 0% extract

5th 2.4 2.6 2.8 2.1 1.6 1.8 2.6 2.3 2.4 2.6 2.9 2.7 1.6 1.7 1.6

7th 2.8 3.0 3.1 3.0 2.8 2.7 2.8 3.0 2.9 3.4 3.2 3.1 2.5 2.4 2.6

Growth in 10% alcoholic extract

5th 1.8 2.4 2.6 1.8 1.5 1.4 1.6 1.8 1.7 2.0 2.0 1.8 1.3 1.4 1.3

7th 2.0 2.8 3.0 2.7 2.5 2.0 2.4 2.5 2.4 2.9 2.8 2.6 2.0 2.0 2.1


5th 1.5 1.6 1.8 1.2 1.1 1.0 1.3 1.4 1.2 1.6 1.4 1.5 1.0 1.1 1.0

7th 1.9 2.2 2.0 2.1 2.2 2.0 1.9 2.0 1.8 2.4 2.3 2.1 1.8 1.7 1.8


5th 1.2 1.3 1.6 1.0 0.8 0.9 1.0 1.2 1.2 1.4 1.2 1.3 0.8 0.9 0.9

7th 1.7 1.3 1.5 1.4 2.0 1.5 1.5 1.4 1.4 1.7 2.0 1.8 1.3 1.4 1.5

DISCUSSION AND CONCLUSION

Studies on herbal plant extracts showed that the

various solvent extracts showed promising

antimicrobial activity against fungal human

pathogens. These extracts can be utilized for

isolation and characterization of therapeutically

active chemical constituents used in modern

medicines. Alcoholic plant extract used here showed

significant antifungal activity against Aspergillus

niger. So this antifungal property provides a

scientific basis for the use of these plants as suitable

antifungal agent. This extract can be used against

infection caused by Aspergillus niger. This study

also encourages that these plant should be cultivated

in large scale to increase the use of these plant in

traditional medicine. Results with different alcoholic

extract concentration of Alstonia scholaris,

Argemone maxicana, Datura alba, Solanum nigrum

and Solanum xanthocarpum on the radial growth of

pathogenic fungus like Aspergillus niger, clearly

shows that alcoholic extract concentration inhibits

radial growth of opportunistic fungi. Result indicates

that inhibition of fungal growth increase with the

increase in the concentration of alcoholic extracts.

REFERENCES

Akerele, O. Summary of WHO (1993). Guidelines

for the assessment of herbal medicines HerbalGram,

28: 13-19.

Antony, M., James, J., Chandra Shekhar Misra,

C.S., Mundur Sagadevan, L.D., (2012). Thaliyil

Veettil, A.K. and Thankamani, V. Anti

mycobacterial activity of the plant extracts of

Alstonia scholaris. International Journal of Current

Pharmaceutical Research, 4(1): 40-42

Cowan, M.M. (1999). Plant products as

antimicrobial agents. Clinical Microbiology Reviews,

12: 564-582

Cragg, G.M., Newman, D.J. and Snader, K.M. (1997). Natural products in drug discovery and

development. Journal of Natural Products, 60: 52-60

George N. (1997). Agrios plant pathology, fifth

edition

Keeler, R.F. and T.U., A.T. (1991). Toxicology of

plant and fungal compounds in: Handbook of

Naturals Toxins, Vol.6; N Y: Marcel Dekker, Inc,

665

Prince, L. and Prabakaran, P. (2011). Antifungal

activity of medicinal plants against plant pathogenic

fungus Colletotrichum falcatum. Asian Journal of

Plant Science and Research, 1 (1): 84-87

Shu, Y.Z. (1998). Recent natural products based

drug development: A pharmaceutical industry

perspective. Journal of Natural Products, 61: 1053-

1071

Thara, K.M. and Zuhra, F. (2013). Biochemical,

HPLC, LC-MS Analysis and Biological activities of

methanol extract of Alstonia scholaris. International

Journal of Phytotherapy, 3 (2): 61-74

Udgirkar, R. F., Kadam, P. and Kale, N. (2012).

Antibacterial activity of some Indian medicinal plant:

a review. International journal of universal

pharmacy and biosciences, 1 (1): 1-8.

412 ARUN KUMAR, VIJAI MALIK AND SHRUTI SAINI



EFFICACY OF BIO-AGENTS AND ORGANIC AMENDMENTS AGAINST

SCLEROTIUM ROLFSII CAUSING COLLAR ROT OF CHICKPEA

Santosh Lahre, N. Khare and Tikendra Kumar*

Department of Plant Pathology, Indira Gandhi Agricultural University,

Raipur 492006, Chhattisgarh, India.



Abstract: Chickpea is cultivated throughout the Chhattisgarh state and mostly grown in kanhar soil in Chhattisgarh plains.

However, chickpea productivity is low due to susceptibility of the crop to different biotic and abiotic stresses. The collar rot

disease of chickpea caused by Sclerotium rolfsii, which is soil borne and fast spreading fungus, causes considerable damage

to the plant stand. The collar rots of chickpea caused by S. rolfsii, can cause considerable loss to plant stand when soil

moisture is high and temperature is warm (nearly 300C ) at sowing time. Drying of plants with foliage turned slightly yellow

before death, scattered throughout the field is an indication of collar rot infection. The study of bio-agent and organic

amendment application revealed that all the treatments significantly increased seed germination and reduced collar rot

incidence. Seed treatment with bio-agent Trichoderma and Neem cake application in soil was found to be the most effective

recording maximum seed germination and minimum mortality followed by Trichoderma with Mustard cake and

Trichoderma with Karanj cake combination under natural condition.

Keywords: Collar rot of chickpea, Sclerotium rolfsii, Trichoderma spp, Bio-agents

INTRODUCTION

hickpea is an important pulse crop grown all

over the world; it occupies the premier position

in terms of area as well as production. In India,

chickpea is grown over 6.93 m ha with the

production of 5.60 m tones. Chickpea contributes

about 37 per cent of the total pulse production in the

country. Chhattisgarh contributes a 0.26 m tone that

is about 4.43 per cent of total chickpea production of

India (Anonymous, 2008). Sclerotium rolfsii causing

collar rot is an important soil borne and fast

spreading fungal pathogen causes considerable

damage to economically important crops like

(chickpea, soybean, groundnut, beans, clover, peas

and lentil). Under field conditions, the S. rolfsii has

been reported to cause 30 to 60 per cent reduction in

yield of chickpea (Prasad, 2005). The collar rots of

chickpea caused by S. rolfsii, can cause considerable

loss to plant stand when soil moisture is high and

temperature is warm (nearly 300C ) at sowing time.

Drying of plants with foliage turned slightly yellow

before death, scattered throughout the field is an

indication of collar rot infection. The disease

generally appear within two weeks of sowing and the

younger plants collapse but older ones turn yellow

and may dry without collapsing. The younger plants

exhibit clear rotting at the collar region. The rotten

portion is often covered with white mycelial strands

of S. rolfsii. Looking towards the above facts an

attempt was made to study the Efficacy of bio-agents

and organic amendments against Sclerotium rolfsii.

MATERIAL AND MATHOD

Seed treatment

Seed treatment was done with Trichoderma @ 4

gm/kg seeds in T1. In treatment T2, T3 and T4 neem

cake, mustard cake and karanj cake respectively was

mixed in the soil and seeds were not treated with

Trichoderma. Treatment T5, T6 and T7 received

seed treatment with Trichoderma and soil treatment

with neem cake, mustard cake and karanj cake

respectively, was done. No seed treatment and soil

treatment served as control.

Seedling inoculation

In seedling stage, test fungus was inoculated at the

collar region of plants in two lines of each plot and

then irrigated lightly by hand shower and mortality

were recorded 15 days after inoculation.

Experimental details

Season : Rabi – 2007

Situation : Upland and Kanhar (Vertisol)

Design : Randomized Block Design

(RBD)

Replications : Three

Treatments : Eight

Variety : JG-315

Plot size : 2 x 3 m2

Seed rate : 80 kg/ha

Date of sowing : 7 November, 2007

C

RESEARCH ARTICLE

414 SANTOSH LAHRE, N. KHARE AND TIKENDRA KUMAR

Details of Treatments

S. N. Treatment Notation

1 Trichoderma spp T1

2 Neem cake T2

3 Mustard cake T3

4 Karanj cake T4

5 Trichoderma spp + Neem cake T5

6 Trichoderma spp + Mustard cake T6

7 Trichoderma spp + Karanj cake T7

8 Control T8

Observation recorded

1. Total plant population per plot

2. Mortality were recorded after 15, 30, and 45 days of sowing.

3. Yield of plots.


Efficacy of bio-agent with organic amendments

against plant mortality caused by S. rolfsii (under

natural condition)

Results (Table1) revealed that all the organic

amendment significantly increased seed germination

and reduced collar rot incidence over untreated

control. However, among all treatments, seed

treatment of bio-agent Trichoderma with Neem cake

soil application was found to be the most effective

recording maximum seed germination (93.05%) and

minimum total mortality (8.32%) followed by

Trichoderma with mustard cake (85.03 & 9.55%)

and Trichoderma with karanj cake (80.86 &

11.41%).

Efficacy of bioagent with organic amendments

against plant mortality caused by S. rolfsii (under

inoculated condition)

After completion of all observations two lines of

each treatment were artificially inoculated by test

fungus Sclerotium rolfsii in all three replications. The

results (Table 2) revealed that among all treatments,

seed treatment of bio-agent Trichoderma and soil

application with Neem cake found to be most

effective with maximum germination (96.29%) and

least total mortality (18.51%), followed by

Trichoderma and Mustard cake (92.59%) and

mortality per cent (25.92) and Trichoderma with

Karanj cake (92.59%) and mortality per cent (25.92).

These treatments were statistically at par with other

for germination and total mortality. Upmanyu et al.

(2002) reported that soil amendment with cotton,

mustard and neem cakes were effective in reducing

pre and post emergence incidence of root rot of

frenchbean under glasshouse and field condition.

Prasad et al. (1999) reported that isolates of

Trichoderma and Gliocladium sp. inhibited mycelial

growth (54.9 to 61.4%) and suppressed the sclerotial

production (31.8 to 97.8%) of S. rolfsii, the causal

organism of root and collar rot of sunflower in-vitro.

Bhoraniya et al. (2003) found that castor oil cakes

reduced the stem rot disease (caused by Sclerotium

rolfsii) incidence of chilli by 78.57 per cent, while

sesame oil cake against foot rot of brinjal (Siddique

et al., 2002). Similarly, neem cake against wilt of

bell pepper (Chowdary et al. 2000) and mustard cake

against wilt of potato (Baker and Khan, 1981) were

superior in controlling the disease caused by S. rolfsii

in pot culture.

Table 1. Effect of bio-agent with organic amendments, on plant mortality under natural conditions

S. No. Treatment Germination* (%) Mortality*(%) Total

mortality

(%)

15 DAS 30 DAS 45 DAS

1

2

3

4

Trichoderma sp.

Neem cake

Mustard cake

Karanj cake

72.99

(58.66)**

66.82

(54.87)

66.97

(55.18)

61.88

2.46

2.77

3.24

3.39

5.55

5.55

5.86

5.40

6.01

5.40

6.17

5.40

14.02

(2.24)***

13.72

(2.23)

15.27

(2.35)

14.19


5

6

7

8

T. sp with neem cake

T. sp with mustard

cake

T. sp with karanj cake

Control

(51.91)

93.05

(74.82)

85.03

(67.80)

80.86

(64.30)

57.40

(49.47)

1.54

2.00

1.85

4.62

3.24

3.54

5.09

9.41

3.54

4.01

4.47

9.25

(2.27)

8.32

(1.79)

9.55

(1.90)

11.41

(2.04)

23.28

(2.84)

SEm +

CD (p=0.05)

0.40

1.22

3.05

8.45

2.74

7.61

3.32

9.20

0.05

0.17

*=Average of three replications

DAS= Days after sowing

**=Data in parenthesis show Arc sine transformation

***=Sq. root transformation value in parenthesis

Table 2. Effect of bio-agent with organic amendments on collar rot under artificially inoculated conditions

S. No. Treatments Germination*

(%)

Mortality*

(%)

Total

mortality

(%)

L1 L2

1

2

3

4

5

6

7

8

Trichoderma sp.

Neem cake

Mustard cake

Karanj cake

T. sp with neem cake

T. sp with mustard cake

T. sp with karanj cake

Control

85.18

(68.67)**

85.18

(67.44)

81.48

(63.72)

74.07

(59.40)

96.29

(77.30)

92.59

(74.51)

92.59

(74.51)

48.15

(43.91)

14.81

18.51

25.92

22.22

7.40

14.81

11.11

33.33

14.81

22.22

29.62

29.62

11.11

11.11

14.81

37.03

29.62

(3.91)***

40.74 (4.56)

55.54

(5.31)

51.86

(5.12)

18.51

(3.10)

25.92

(3.66)

25.92

(3.66)

70.37

(5.97)

SEm +

CD (p=0.05)

2.34

7.12

0.20

0.56

0.60

1.67

0.19

0.65

*= Average of three replications

**=Data in parenthesis show Arc sine transformation

416 SANTOSH LAHRE, N. KHARE AND TIKENDRA KUMAR

***=Sq. root transformation value in parenthesis

L1= Line no. 1

L2= Line no. 2

REFERENCES

Anonymous, (2008). Project Coordinator`s Report

AICRP on Chikpea IIPR, Kanpur Ann. 21-22

Baker,A. and Khan, A. A. (1981). Effect of

nitrogenous amendments on the incidence of

Sclerotium wilt of potato. Potato Res. 24(3): 363-

365.

Bhoraniya, M. F., Khandar, R. R. and Khunti, J.

P. (2003). Integrated management of stem rot of

chilli with fungicides, oil cakes and biocontrol agent

Trichoderma harzianum. Adv. Pl. Sci. 16(1): 59-61.

Chowdary, A. K., Reddy, D. R., Rao, K. C.,

Reddy, T. B. and Reddy, I. P. (2000). Integrated

management of Sclerotium wilt disease of bell

pepper (Capsicum annum L.) Indian J. Pl. Prot.

28(1): 15-18.

Prasad, R. D., Rangeshwaran, R. and Kumar, P.

S. (1999). Biological control of root and collar rot of

sunflower. Indian J. Mycol. Pl. Pathol. 29(2): 184-

188.

Prasad, R. D. (2005). Status of biological control of

soil born plant pathogens. Indian J. Agri. Sci. 77(9):

583-588.

Siddique, M. A. B., Meah, M. B., Siddqua, M. K.,

Haque, M. M. and Babar, H. M. (2002). Control of

foot rot of brinjal using fungicide and organic

amendment. Bangladesh J. Pl. Pathol. 18(1): 61-64.

Upmanyu, S., Gupta, S. K. and Shyam, K. R. (2002). Innovative approaches for the management

of root rot and web blight of Frenchbean. J. Mycol.

Pl. Pathol. 32(3): 317-331.



INSECT PESTS COMPLEX ASSOCIATED WITH BASMATI RICE WITH

WESTERN PLAIN ZONE OF UTTAR PRADESH, INDIA

Kaushlendra Kumar*, S.K. Sachan and D.V. Singh

Department of Entomology

Sardar Vallabhbhai Patel University of Agriculture and Technology, Meerut-250110 (U. P.)


Abstract: Insect pests complex associated with basmati rice were studied during Kharif, 2014 and 2015 at Crop Research

Center of Sardar Vallabhbhai Patel University of Agriculture and Technology, Meerut. During the study period, fifteen

insect species were encountered on basmati rice in western plain zone of Uttar Pradesh which belong to 7 orders viz.

lepidoptera (yellow stem borer, leaf folder, striped rice stem borer, rice case worm and swarming caterpillar), homoptera

(green leaf hopper, brown plant hopper, and white backed plant hopper), heteroptera (rice gundhi bug), hetroptera (rice

mealy bug), coleoptera (rice root weevil and white grub), isoptera (termite) and orthoptera (Kharif grass hopper and grass

hopper).

Keywords: Insect pests, Basmati rice, Grass hopper

INTRODUCTION

ice is the major food of the largest population of

the world. About 90 per cent rice in the world is

grown and consumed by the population of the Asian

countries (Samanta, 2014). In India, it occupies an

area of about 43.95 million hectare with total

production of 106.54 million tones and productivity

of 2.4 tones per hectare (Anonymous 2014). Basmati

rice crop suffers severely due to attack of various

insect pests which reduces its yield and quality. More

than 300 species of insects have been reported to

attacked rice crop from the germination of nursery

till its harvests (Jadhao and Khurad, 2011). In

general, yield loss due to insect pest of rice has been

estimated at about 25 per cent in different rice

ecosystem (Sachan et al., 2006 and Dhaliwal et al.,

2010). Keeping in view this fact, the present

investigation to update the information about insect

pests complex associated with basmati rice in this

region.

MATERIAL AND METHOD

An experiment were conduct during two consecutive

crop season, viz Kharif, 2014 and 2015 at Crop

Research Centre, Sardar Vallabhbhai Patel

University of Agriculture and Technology, Meerut

(U. P.). The rice cultivar PB-1 was sown during mid

of June and transplanted in the second week of July

in both the years and adopted recommended practices

to rice good crop. Observations on insect pests

associated with rice were recorded from the

germination of seedlings till the harvest of crop at

weekly interval. The insect were collected and

identified.


During the period of study, 15 insect species

belonging to 7 orders were recorded on basmati rice

at different crop growth stage. Among them, yellow

stem borer, Scirpophaga incertulas Walker and leaf

folder, Cnaphalocrocis medinalis Guenee were

found as major pests. The brown plant hopper,

Nilaparvata lugens Stal., white grub, Holotrichia

consanguinea Blanch, termite, Odontotermes obesus

Romb. and Kharif grass hopper, Hieroglyphus

banian Fab. were found moderately damaging the

crop. The rice swarming caterpillar (army worm),

Spodoptera mauritia Boisduval, striped rice stem

borer, Chilo suppressalis (Walker), rice case worm,

Nymphula depunctalis (Guenee), green leaf hopper,

Nephotettix virescens Distant, white backed plant

hopper, Sogatella furcifera Horvath, rice gundhi bug,

Leptocorisa acuta Thunb, rice root weevil,

Echinocnemus oryzae Marshall, rice mealy bug,

Brevennia rehi (Lind.) and grass hopper, Oxya

fuscovittata Marshall recorded on the crop were of

less importance and extent of their damage was

found without much economic loss (Table 1).

Yellow stem borer is the most destructive and

dominant species of rice crop in this region and is

reported to occur throughout the country and other

Asian countries. The appearance of pest started in the

beginning of July and remained active throughout the

crop season. The newly hatched larvae enter in leaf

sheath and bore into the stem near the node and

causing death of central shoot „dead hearts‟ in

vegetative stage and „white ear head‟ at milky stage,

respectively. This resulted in chaffy grains. The

severity of this pest was recorded from beginning of

August to September end. The damage of yellow

stem borer on rice crop has also been reported by

Kumar and patil (2004), Dogra and Amit (2005),

Satpathi et al. (2012) and Gangwar et al. (2015).

Sachan et al. (2006) also reported the severe

R

RESEARCH ARTICLE

418 KAUSHLENDRA KUMAR, S.K. SACHAN AND D.V. SINGH

incidence of S. incertulas on basmati rice throughout

the crop season in tarai region of Uttar Pradesh.

Rice leaf folder is another major pest of basmati rice

recorded from middle of August to September end. It

has inflicted severe losses on rice crop. The newly

hatched larvae crawl to the base of the youngest

unopened leaf and begin to feed and then migrate to

an older leaf and folds leaf together. Under heavy

infestation reduced the general vigor and

photosynthetic ability of an infested plant.

Considerable losses to paddy crop due to leaf folder

have been reported by several workers like Sachan et

al. (2006), Jadhao and Khurad (2011) and Saini et al.

(2015).

The striped stem borer is a polyphagous pest

frequently occurring in the rice fields of almost all

fields in the area. The first symptoms of damage

were the drying off of growing points and the

surrounding leaves and finally, leaves fall off. In the

infested field white heads stand erect and contain

empty and unfilled glumes. The striped yellow stem

borer on rice crop has also been reported by Pathak

(1968), Singh and Singh (2014) and Gangwar et al.

(2015) reported that important insect pest of rice in

India and South- East Asia.

The swarming caterpillar is considered to be a

sporadic pest which occasionally causes serious

losses to upland rice crop. Larvae of this pest fed on

the upper portion of rice canopy by defoliating leaves

during night. This insect recorded as minor pest in

the month of July –August. This swarming caterpillar

on rice crop has also been reported by several

workers like Jadhao and Khurad (2011), and

Gangwar et al. (2015).

The rice case worm is an important insect pest of

rice. The damaging stage is the larvae that live in

sections of leaves cut from young rice plants and

rolled tubes called cases. Rice at seedling and

tillering stages are the preferred host but does not

occur after maximum tillering. The insect was

recorded as minor pest of this region. The damage of

this insect has also been reported by Dale (1994),

Singh and Singh (2014) and Gangwar et al. (2015).

This pest is widely distributed in rice growing

countries of Asia.

The incidence of green leaf hopper, Nephotettix

virescens (Distant) was recorded in August-

September. Both, nymphs and adults of this insect

sucked sap from the leaves and tender parts of plants

by turning them yellow. Brown plant hopper,

Nilaparvata lugens (Stal) is another important pest of

rice. Its infestation was recorded from middle of

August to September end. As a result of feeding by

both nymphs and adults at the base of the tillers,

plants turns yellow and dry up rapidly. At early

infestation round yellow patches appeared which

soon turn brownish due to the drying up of the plants.

This condition is called “hopper burn”. Complete

destruction of the crop was recorded in severe cases.

White backed plant hopper, Sogatekka furcifera

(Horvath) was recorded in August - September as

minor pest in this region. It sucked the sap from

tender leaves, thus causing yellowishness of them.

The honeydew produced by the hoppers serves as a

medium for growth of sooty mould.

The damage caused by green leaf hopper, brown

plant hopper, and white backed plant hopper on rice

crop has been reported by various workers such as

Sachan et al. (2006) and Singh and Singh (2014).

However the finding of Atwal and Dhaliwal (2005),

Srivastava (2006), Sachan et al. (2006), Singh and

Singh, (2014) and Sharma (2015) are also in

conformity with present finding that green leaf

hopper and brown plant hopper were the pest of

basmati rice.

The rice gundhi bug was recorded as important pest

of rice crop in this region. Both nymphs and adults

sucked the sap of developing grains during milky

stage and thus make them chaffy. Whole panicle

becomes white colored (chaffy) under severe

infestation. Its occurrence was recorded during

September–October. Atwal and Dhaliwal (2005),

Sachan et al. (2006), Kashyap (2013), Singh and

Singh, (2014), Gangwar et al. (2015) and Sharma

(2015) also reported the damage of this pest during

September-October on rice crop.

Rice mealy bug is a polyphagous pest. The mealy

bug occurs in colonies attached to the stem and leaf

sheaths of plants. They sucked the sap from the plant.

The high incidence of this pests inhibits panicle

emergence and plant may even dry. This pest first

time reported in this region. Singh and Singh (2014)

has been reported incidence of rice mealy bug in

northeastern Uttar Pradesh.

Rice root weevil was also recorded a common pest of

basmati rice at vegetative stages. Grubs fed on the

roots and rootlets of young rice plants, resulted in

stunting and non formation of tillers. The leaves turn

yellow and develop a rusty appearance and the plants

eventually die. Incidence of this pest was noticed in

the month of July and August. Singh and Singh

(2014) and Gangwar et al. (2015) also reported the

occurrence of this pest on rice crop.

White grubs are a serious polyphagous pest and

damage almost all the Kharif crops from June to

October. The young larvae feed upon roots and

decaying vegetation throughout the summer and, in

autumn, migrate downward (to a depth of up to 1.5

meters) and remain inactive until the following

spring. The severe damage occurs as the larvae move

near the soil surface to feed on the roots of the plants.

The beetles overwinter in the soil, emerging the

following year in May or June when feeding, mating,

and egg-laying take place. The insect was recorded

from July to October i.e. throughout the crop season.

Atwal and Dhaliwal (2005), Kashyap (2013) and

Saini (2015) also reported the incidence of this insect

from July to October on various Kharif crops.

Termite is a polyphagous social insect, also caused

damage to the rice crop by feeding on the roots of the


plants. The growing shoots withered and died. The

damaged plants pulled out easily. The incidence of

this pest was observed throughout the crop season.

Such type of damage has earlier reported on different

crops by Atwal and Dhaliwal (2005), Prasad and

Prasad (2006), Sachan et al. (2006), Srivastava

(2006), Kashyap (2013), Singh and Singh (2014)

and Saini (2015).

The two species of grasshoppers namely

Hieroglyphous banian (Fab.) and Oxya fuscovittatta

(Marshall) were found to attack basmati rice in this

region. Nymphs and adults of grasshoppers fed on

the leaves by making holes and were found active

throughout the crop season while latter was found in

the month of August and September as minor pest.

Both these species of grasshopper are polyphagous

pest and have earlier been reported by Sachan et al.

(2006), Prasad and Prasad (2006), Kashyap (2103),

Singh and Singh (2014), Sharma (2015), Gangwar et

al. (2015) and Saini (2015) on paddy crop. Usmani et

al. (2012) observed Kharif grass hopper as a pest of

paddy from central Uttar Pradesh.

Table 1. Insect pests complex associated with basmati rice during kharif 2014 and 2015 Common Name Scientific Name Order - Family Damaging stage of

the pest

Severity of

the pests

Yellow stem borer Scirpophaga incertulas (Walker)

Lepidoptera : Pyralidae Larvae Severe

Striped rice stem borer

Chilo suppressalis (Walker) Lepidoptera : Pyralidae Larvae Low

Leaf folder Cnaphalocrosis medinIalis

(Guenee)

Lepidoptera : Pyralidae Larvae Severe

Swarming caterpillar

(Army worm)

Spdoptera mauritia (Boisduval) Lepidoptera :Noctuidae Larvae Low

Rice case worm Nymphula depunctalis (Guenee) Lepidoptera: Pyralidae Larvae Low

Green leaf hopper Nephotettex virescens (Distant) Homoptera :Cecadellidae Nymphs and adults Low

Brown plant hopper Nilaparvata lugens (Stal) Homoptera : Delphacidae Nymphs and adults Moderate

White Backed plant

hopper

Sogatekka furcifera (Horvath) Homoptera : Delphacidae Nymphs and adults Low

Rice gundhi bug Leptocorisa acuta (Thumb) Hetroptera :Coreidae Nymphs and adults Low

Rice mealy bug Brevennia rehi (Lind.) Hemiptera:Pseudococcidae Nymphs Low

Rice Root Weevil Echinocnemus oryzae (Marshall)

Coleoptera : Curculionidae Grubs and adults Low

White grub Holotrichia consanguinea

(Blanch)

Coleoptera : Curculionidae Grubs and adults Moderate

Termite Odontotermes obesus (Romb) Isoptera : Termitdae Worker Moderate

Kharif grass hopper Hieroglyphus banian (Fab) Orthoptera : Acrididae Nymphs and adults Moderate

Grass hopper Oxya fuscovittata (Marshall) Orthoptera : Acrididae Nymphs and adults Low

REFERENCES

Anonymous, (2014). Ministry of Agriculture,

Government of India. http:// www.Indiastattictic.

Com.

Dhaliwal,G.S., Jindal,V. and Dhawan, A. K., (2010). Insect pest problems and crop losses:

changing trends. Indian. J. Ecol., 37, 1-7.

Gangwar, R. K., Javeria, S., Yadav, K., Tyagi, S.

And Singh, R. (2015). Survey and surveillance of

major insect-pests of basmati rice in western Uttar

Pradesh (India). International Journal of Research in

Applied, Natural and Social Science, 3 (3): 1-8.

Kashyap, A. K. (2013). Evaluation of some novel

insecticides against yellow stem borer, Scripophaga

incertulas (Walker) and rice leaf folder,

Cnaphalocrocis medinalis (Guenee) and their effect

on natural enemies in basmati rice. M. Sc (Ag)

Thesis, S. V. P. Univ. of Agric & Tech. Meerut.

Kumar, V. and Patil, B. V. (2004). Insect pest fauna

to rice in Tungabhadra Project area of Karnataka,

420 KAUSHLENDRA KUMAR, S.K. SACHAN AND D.V. SINGH

during Kharif season. Karnatka J. Agric. Sci., 17 (3):

580-581.

Pathak, M. D. (1968). Ecology of rice pests, Annu.

Rev. Entomol., 3: 257–294.

Prasad, R. and Prasad, D. (2006). Account of

insect pest problem in rice ecosystem in Ranchi.

Indian J. Ent., 68 (3): 240-246.

Sachan, S. K., Singh, D. V. and Chaudhary, A. S. (2006). seasonal abundance of insect pests associated

with basmati rice. Ann. Pl. Protec. Sci., 14 (1):218-

220.

Saini, U. P., Sachan, S. K., Singh, A. P. B. and

Kumar, K, (2015). Insect pests associated with

basmati rice in western plain zone of Uttar Pradesh,

India. Plant Archives, 15 (2): 775-777.

Samanta, A., Chakraborti, K., Alam, S. K. F.,

Das, B. C. and Patra, S. (2014). Pest surveillance in

lcc and non-lcc rice plots by participatory rural folk

appraisal. The Ecoscan, 8(3&4): 211-213.

Satpathi, C. R., Chakraborty, K., Shikari, D. and

Acharjee, P. (2012). Consequences of feeding by

yellow stem borer on rice cultivar swarna mashuri.

World Appl. Sci. J., 17: 532-539.

Sharma, R. (2015). Evaluation of microbial and

chemical insecticides against yellow stem borer,

Scripophaga incertulas (Walker) and leaf folder,

Cnaphalocrocis medinalis (Guence) of basmati rice.

M. Sc (Ag) Thesis, S. V. P. Univ. of Agric & Tech.

Meerut.

Singh, B. B. and Singh R. (2014). Major rice insect

pests in north eastern Uttar Pradesh. Int. J. Life Sci.

Biotech. & Pharma. Res., 3(1): 124-143.

Usmani, M. K., Nayeem, M. R. and Akhtar, M. H. (2012). Field observations on the incidence of

grasshopper fauna as a pest of paddy and pulses. Eur.

J. Exp. Biol., 2: 1912-1917.



PLANTS AS A SOURCE OF DIURETIC ACTIVITY AND STUDY OF 3-(6-

ARYLIMIDAZO[2,1-B]THIAZOL-3-YL)-2-METHYLCHROMONE SYSTEM AS

DIURETIC AGENT

Vinay Prabha Sharma*

Department of Chemistry ; Meerut College , Meerut – U.P. (India)



Abstract: Diuretic agents increase urine volume and are effective in heart failure , renal failure and maintain Na+ ion

balance. They are also effective in hypertension and nephrosis. Though plants possess diuretic activity , but their delayed

action needs to use quickly acting agents . In this paper study on 3-(6-Arylimidazo[2,1-b]thiazol-3-yl)-2-methylchromones

as diuretic agents is being discussed . Lead for diuretic activity has been found in this system .

Keywords: Diuretic activity, Chromones, Lead, Structure activity relationship (SAR)

INTRODUCTION

iuretics are the drugs that increase urine volume

as well as flow and clinically useful diuretics

increase the rate of excretion of Na+ ions (natriuresis)

and accompanying ions like Cl- ions (Jackson, 2001)

. They adjust urine volume and composition of body

fluids in situations like hypertension, heart failure,

renal failure , nephrosis etc. (Jackson, 2001) . Most

of the diuretics have adverse effects like fatigue,

weakness and impotence. Plant based diuretics

include caffeine present in tea, coffee and cola which

inhibit sodium ion re-absorption . Most of the

diuretics are effective in promoting sodium ion

excretion; but all cause potassium ion loss

(Vanamala et al. , 2012) . Moreover, resistance

develops to diuretics (Brater, 1983). More than 650

herbal preparations in the form of tablets, decoctions,

tinctures etc. have shown diuretic activity (Chopra et

al., 1986). Achyranthes aspera Linn .also possess

diuretic activity (Srivastava et al., 2011). Aqueous

extract of mango bark ( Mangifera indica ) is also

diuretic (Shree Devi , 2011). Kane et al. (2009)

reported potentiation of diuretic activity through

ethanolic extract of Euphorbia thymifolia. Thus,

various plant parts are good diuretic agents (Dutta et

al. , 2014) ; but their action is delayed . Therefore,

there is need to develop other diuretics which are

non-resistant and non-toxic. Probably diuretic

activity in plants is due to the presence of flavonoids

and alkaloids in them (Vanamala et al., 2012) . Few

chromone derivatives have shown diuretic activity in

past (Sharma, 2015). Hence, it was thought to study

diuretic activity in 3-(6-Arylimidazo[2,1-b]thiazol-3-

yl)-2-methylchromones .

MATERIAL AND METHOD

3-(6-Arylimidazo[2,1-b]thiazol-3-yl)-2-

methylchromones were synthesized by author in the

Department of chemistry , Kurukshetra University ,

Kurukshetra ( Garg et al. , 1985 ; Sharma , 2005) .

The diuretic activity was tested at CDRI , Lucknow .

Activity of compounds was compared with

Chlorothiazide standard (value for which is taken as

100) [ Table-1] .


6-Chloro-3-(6-phenylimidazo[2,1-b]thiazol-3-yl)-2-

methylchromone (VPS-10) showed diuretic activity

equal to 100 which is same as for chlorothiazide

standard . However, compound is a bit toxic with

ALD50 = 681 . VPS-11 with two halogens (chlorine

atoms) , one at C6 of chromone ring and other at p-

position of phenyl ring present at C6- position of

imidazothiazolyl moiety , shows decrease in activity

which becomes equal to 93 . This compound

becomes safer [ALD50 > 1000] . Here one additional

chlorine atom at conjugated position of phenyl ring

decreases activity and toxicity as well. VPS-12 has

one additional methyl group compared to VPS-10 at

C7- position of chromone ring; it exhibited less

activity than VPS-10. It is inferred that additional

methyl groups result in reduction of diuretic activity

( c. f. Gupta , 2014) . One additional methyl group

also reduced toxicity as ALD50 changes from 681 to

1000.

VPS-13 has two halogen atoms , Cl at C6- position of

chromone moiety and Br at p- position of phenyl ring

present at C6 – of imidazothiazolyl moiety . This

compound also possesses a methyl group at C7 – of

chromone ring. Here reduction in diuretic activity is

due to substituent Br and methyl group . Therefore ,

activity is much reduced and becomes equal to 68

because of the additive effect of Br and CH3 .

Toxicity of this compound is low because both Br

and CH3 decrease toxicity. 3-(6-(p)-

chlorophenylimidazo[2,1-b]thiazol-3-yl)-2,6-

dimethylchromome (VPS-14) has intermediate

activity of 87 as methyl group decreases the activity

and Cl substituent increases the activity in

comparison to methyl group. As both these

D

SHORT COMMUNICATION


422 VINAY PRABHA SHARMA

substituent decrease the toxicity the compound is

safer for use with ALD50 value of 1000.

A look at these activities shows that additional

halogen atom either Cl or Br decreases activity to

same extent [activity of VPS-10 > VPS-11 by 7-units

and activity of VPS-12 is > VPS-13 by 8-units]

which may be ascribed to their same electronic

effects (Silverman , 2004). Replacement of Cl by

CH3 group decreases the activity by 6-units [ c.f.

activity of VPS-11 and VPS-14].

Table 1. Effect of substitution on the diuretic activities of 3-(6-arylimidazo[2,1-b]thiazol-3-yl)-2-

methylchromones :

S.No. Code Name of the compound Activity ALD50

1. VPS-10 6-chloro-3-(6-phenylimidazo[2,1-b]thiazol-3-yl)-2-methylchromone 100 681

2. VPS-11 6-chloro-3-(6-(p)-chlorophenylimidazo[2,1-b]thiazol-3-yl)-2-

methylchromone

93 >1000

3. VPS-12 6-chloro-3-(6-phenylimidazo[2,1-b]thiazol-3-yl)-2,7-

dimethylchromone

76 1000

4. VPS-13 6-chloro-3-(6-(p)-bromophenylimidazo[2,1-b]thiazol-3-yl)-2,7-

dimethylchromone

68 1000

5. VPS-14 3-(6-(p)-chlorophenylimidazo[2,1-b]thiazol-3-yl)-2,6-

dimethylchromone

87 1000

6. Standard Chlorothiazide 100 Drug

CONCLUSION

Methyl group at C6 and C7 – positions of chromone

ring decreases the activity and introduction of

halogen atom at p- position of aryl group present at

6-position of imidazothiazolyl moiety of 3-(6-

arylimidazo[2,1-b]thiazol-3-yl)-2-methylchromones

though decreases the activity but increases the

diuretic activity in comparison to methyl group

which may be attributed to their electronic effects .

As all the tested 3-(6-arylimidazo[2,1-b]thiazol-3-

yl)-2-methylchromones exhibited good diuretic

activity so this system is a lead for this activity .

ACKNOWLEDGEMENT

Facilities provided by Kurukshetra University ,

Kurukshetra and CDRI-Lucknow are thankfully

acknowledged.

REFERENCES

Brater, D.C. (1983). Resistance to loop diuretics.

Drugs , 30 : 427-443.

Chopra, R.N.; Nayer, S.L. and Chopra, L.C. (1986). Glossary of Indian Medicinal Plants

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